Modification of the length and structure of the linker of N6-benzyladenosine modulates its selective antiviral activity against enterovirus 71

Article abstract of DOI:10.1016/j.ejmech.2016.01.036

Very recently, we demonstrated that N⁶-isopentenyladenosine, a cytokinin nucleoside, exerts a potent and selective antiviral effect on the replication of human enterovirus 71. The present study is devoted to the structure optimization of another natural compound: N⁶-benzyladenosine. We mainly focused on the exploration of the size and nature of the linker between the adenine and the phenyl ring, as well as on the necessity of the D-ribose residue. More than 30 analogues of N⁶-benzyladenosine were prepared and their antiviral properties were evaluated. Two main methodologies were used for preparation: N⁶-acetyl-2′,3′,5′-tri-O-acetyladenosine can be regioselectively alkylated either by alkyl halides under base promoted conditions or by alcohols in Mitsunobu reactions. After deacylation with 4 M PrNH₂ in MeOH at room temperature for one day, the desired products were obtained in overall high yields. Analysis of the structure-activity relationship clearly shows that the optimal size of the linker is limited to 2 or 3 atoms (compounds 4-7). 2′-Deoxyadenosine derivatives did not elicit any inhibitory or cytotoxic effect, while 5′-deoxynucleosides still induced some cell protective antiviral activity. Based on these observations, it can be hypothesized that there may be another mechanism that is at the base of the antiviral activity of these compounds against enterovirus 71 besides a possible 5′-triphosphorylation followed by a putative inhibitory effect on RNA synthesis.

Full text of DOI:10.1016/j.ejmech.2016.01.036

Accepted Manuscript
6
Modification of the length and structure of the linker of N -benzyladenosine
modulates its selective antiviral activity against enterovirus 71
Mikhail S. Drenichev, Vladimir E. Oslovsky, Liang Sun, Aloys Tijsma, Nikolay
N. Kurochkin, Vitali I. Tararov, Alexander O. Chizhov, Johan Neyts, Christophe
Pannecouque, Pieter Leyssen, Sergey N. Mikhailov
PII:
S0223-5234(16)30036-8
10.1016/j.ejmech.2016.01.036
EJMECH 8326
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 25 November 2015
Revised Date: 18 January 2016
Accepted Date: 19 January 2016
Please cite this article as: M.S. Drenichev, V.E. Oslovsky, L. Sun, A. Tijsma, N.N. Kurochkin, V.I.
Tararov, A.O. Chizhov, J. Neyts, C. Pannecouque, P. Leyssen, S.N. Mikhailov, Modification of the
6
length and structure of the linker of N -benzyladenosine modulates its selective antiviral activity against
enterovirus 71, European Journal of Medicinal Chemistry (2016), doi: 10.1016/j.ejmech.2016.01.036.
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to
our customers we are providing this early version of the manuscript. The manuscript will undergo
copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please
note that during the production process errors may be discovered which could affect the content, and all
legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT
6
Modification of the length and structure of the linker of N -benzyladenosine modulates its selective
antiviral activity against enterovirus 71
a1
a1
b
b
a
Mikhail S. Drenichev , Vladimir E. Oslovsky , Liang Sun , Aloys Tijsma , Nikolay N. Kurochkin ,
a
c
b
b
b
Vitali I. Tararov , Alexander O. Chizhov , Johan Neyts , Christophe Pannecouque , Pieter Leyssen ,
Sergey N. Mikhailov *
a
a
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov Str. 32, Moscow
1
19991, Russian Federation
KU Leuven - University of Leuven, Department of Microbiology and Immunology, Laboratory for
b
Virology and Chemotherapy, Rega Institute for Medical Research, Minderbroedersstraat 10, 3000
Leuven, Belgium
c Zelinsky Institute of Organic Chemistry, Russian Academy of Science, Leninsky pr., 47, Moscow
1
19991, Russian Federation
1
Both authors contributed equally.
GRAPHICAL ABSTRACT

ACCEPTED MANUSCRIPT
6
Modification of the length and structure of the linker of N -benzyladenosine modulates its selective
antiviral activity against enterovirus 71
a1
a1
b
b
a
Mikhail S. Drenichev , Vladimir E. Oslovsky , Liang Sun , Aloys Tijsma , Nikolay N. Kurochkin ,
a
c
b
b
b
Vitali I. Tararov , Alexander O. Chizhov , Johan Neyts , Christophe Pannecouque , Pieter Leyssen ,
a
Sergey N. Mikhailov *
a
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov Str. 32, Moscow
1
19991, Russian Federation
b
KU Leuven - University of Leuven, Department of Microbiology and Immunology, Laboratory for
Virology and Chemotherapy, Rega Institute for Medical Research, Minderbroedersstraat 10, 3000
Leuven, Belgium
c Zelinsky Institute of Organic Chemistry, Russian Academy of Science, Leninsky pr., 47, Moscow
1
19991, Russian Federation
1
Both authors contributed equally.
Abstract
6
Very recently, we demonstrated that N -isopentenyladenosine, a cytokinin nucleoside, exerts a potent and
selective antiviral effect on the replication of human enterovirus 71. The present study is devoted to the
6
structure optimization of another natural compound: N -benzyladenosine. We mainly focused on the
exploration of the size and nature of the linker between the adenine and the phenyl ring, as well as on the
6
necessity of the D-ribose residue. More than 30 analogues of N -benzyladenosine were prepared and their
6
antiviral properties were evaluated. Two main methodologies were used for preparation: N -acetyl-
2
′,3′,5′-tri-O-acetyladenosine can be regioselectively alkylated either by alkyl halides under base
promoted conditions or by alcohols in Mitsunobu reactions. After deacylation with 4M PrNH in MeOH
2
at room temperature for one day, the desired products were obtained in overall high yields. Analysis of
the structure-activity relationship clearly shows that the optimal size of the linker is limited to 2 or 3
atoms (compounds 4-7). 2′-Deoxyadenosine derivatives did not elicit any inhibitory or cytotoxic effect,
while 5′-deoxynucleosides still induced some cell protective antiviral activity. Based on these
observations, it can be hypothesized that there may be another mechanism that is at the base of the
antiviral activity of these compounds against enterovirus 71 besides a possible 5′-triphosphorylation
followed by a putative inhibitory effect on RNA synthesis.
1

ACCEPTED MANUSCRIPT
1
. Introduction
Enterovirus 71 (EV71) is a non-enveloped, single-stranded, positive-sense RNA virus that belongs to the
Picornaviridae family. The virus is, together with Coxsackievirus A16 (CV-A16), a major causative
agent of hand-, foot-, and mouth disease (HFMD), a mild and self-limiting illness that mostly affects
children under the age of 5. However, in some patients, the virus can cause severe, potentially lethal
complications such as aseptic meningitis, encephalitis, pulmonary edema, and viral myocarditis [1, 2]. In
recent years, EV71 has caused large outbreaks of HFMD in Asia, during which an increased number of
severe neurological complications, such as encephalitis and acute flaccid paralysis, have been observed
[3]. Furthermore, in the past ten years, EV71 infections have also been reported in countries in America
and Europe [4].
Even though several promising EV71 vaccine candidates are currently in clinical trial [5], to date, there is
still no approved vaccine available for prophylaxis [6, 7, 8]. In addition, it may also be important to have
effective antivirals at hand for the treatment of infected patients with severe disease [9, 10, 11]. At the
moment there are still no antivirals on the market for the treatment of enterovirus infection.
In 2008, Arita and coworkers identified 4 inhibitors of EV71 replication during a screen of the
1
280
6
LOPAC
drug library (Sigma-Aldrich) [12]. Two of these compounds were nucleoside analogs, i.e. N -
6
benzyladenosine (23) and N -(o-methylbenzyl)adenosine (metrifudil). In 2014, Shang and colleagues
6
reported on the anti-EV71 activity of several natural and synthetic nucleosides. In particular N -
benzyladenosine and 2′-ethynyladenosine were shown to have significant antiviral activity against EV71
with an EC of 5 µM and 0.625 µM, respectively [13]. Recently, our research groups demonstrated that
5
0
6
N -isopenthenyladenosine, a natural plant cytokinin (31), also inhibits the replication of EV71 with an
EC of 1.0 ± 0.2 ꢀM [14]. Natural products have been the major sources of drugs in medicinal chemistry.
50
An approach which is based on the structure of natural compounds is rather popular and fruitful [15, 16,
7]. For example, about 100 nucleoside-based drugs have been developed and nucleoside library is a
promising pool for the fishing of new biologically active compounds [18, 19].
1
6
In the present study, we report on the exploration of the properties of the linker structure and length of N -
benzyladenosine-based compounds in an attempt to obtain more potent and selective inhibitors of the
replication of EV71.
2
. Results and discussion
2
.1. Chemistry
2

ACCEPTED MANUSCRIPT
6
Recently, we have developed a new useful and versatile approach for the preparation of N -adenosine
6
6
derivatives by regioselective alkylation of N -alkylation of N -acetyl-2′,3′,5′-tri-O-acetyladenosine (1)
[20] with alcohols under Mitsunobu conditions or with alkyl halides promoted by a base (Scheme 1). The
6
traditional approach for the preparation of N -alkylated adenosines is the substitution of the chlorine atom
in commercially available 6-chloropurine riboside with alkylamines [21, 22] (Scheme 2). The main
6
advantage of our method is the possibility to use both alkyl halides and alcohols for N -modification. This
is important, especially in the case when an amine is not stable or hardly available. The only difficulty
was associated with purification of the final nucleosides from the by-products caused by the Mitsunobu
reaction and acetamide forming under ammonolysis conditions. We concluded that, for this class of
compounds, the method of choice was deacetylation with 4M PrNH in MeOH, which significantly
2
facilitated chromatographic purification [14].
6
The developed protocol for N -alkylation was used successfully for the preparation of isomeric
deoxyadenosine derivatives starting from triacetyl-2′-deoxyadenosine 2 and triacetyl-5′-deoxyadenosine
3
. The key intermediate 3 was prepared by radical Bu SnH reduction of corresponding 5′-chloro-5′-
3
deoxyadenosine derivative in a way similar to 5′-deoxyuridine synthesis [23].
6
6
6
Scheme 1. Synthesis of N -alkyladenosines, N -alkyl-2′-deoxyadenosines and N -alkyl-5′-deoxyadenosines. Reagents and
o
o
conditions: (i) ROH, Ph P, DEAD, THF, 20 C, 24-48 h; (ii) RBr, DBU, CH CN, r.t., 1-3 days; (iii) 4M PrNH in MeOH, 20 C,
3
3
2
2
4 h. (The structure of R is given in Tables 1, 3, 4).
6
6
The preparation of a series of N -aryladenosines and O -alkylinosines (Scheme 2) was started with
2
′,3′,5′-tri-O-acetyl-6-chloropurineriboside (18). The substitution of chloride with various nucleophiles in
unprotected 6-chloropurineriboside is well-known [21, 22]. 6-Chloropurineriboside can be prepared by
deacetylation of 18 [24]. To simplify the procedure, we used compound 18 directly in the substitution
reactions. Interestingly, acetyl groups are completely preserved in the reaction with aniline and the
protected intermediate can be isolated and characterized. The removal of acetyl groups by ammonolysis
affords nucleoside 19. This is quite opposite to the reactions with alkylamines and alcoholates, in which
the complete deacetylation has been observed. The isolation of inosine derivatives 20-22 was performed
3

ACCEPTED MANUSCRIPT
by column chromatography (silica gel) of the whole reaction mixtures after neutralization with neat acetic
acid.
6
6
Scheme 2. Synthesis of N -alkyladenosines and O -substituted inosine derivatives by substitution of chlorine atom in 2′,3′,5′-
o
tri-O-acetyl-6-chloropurineriboside (18). Reagents and conditions: (i) RNH , n-BuOH, DIPEA, 80 C; (ii) 7M NH /MeOH, r.t.,
2
3
o
2
4 h; (iii) DMSO, t-BuOK, ROH, 20 C, 24 h. (The structure of R is given in Tables 1-4).
The structure of compounds was confirmed by NMR spectral data. The distinguishing feature of
unprotected adenosines and their parent bases is the significant broadening of signals of the CH group
2
6
6
attached to the N -atom. Such effect is general for N -substituted nucleosides and their bases. The
possible explanation can be found in the literature and is ascribed to ‘a chemical exchange process
connected to the restricted conformational change’ [25, 26]. Noteworthy, broadening is observed for the
6
6
same compounds when a protective N -acyl group is present and for corresponding O -inosine
derivatives.
2
.2. Biological activity
6
N -Substituted adenines (cytokinins) represent an important group of phytohormones that are involved in
numerous biochemical processes in plants, stimulating cell division and regulating plant growth. In
6
nature, cytokinins can be found with an aromatic side chain (N -benzyladenine and its hydroxylated
6
derivatives – topolins), with an isoprenoid side chain, such as N -(ꢁ2-isopentenyl)adenine (iP), and as
their hydroxylated derivatives – zeatins. The biosynthesis is only well understood in the case of iP.
Cytokinins can occur in nature in the form of free bases, nucleosides (N-glycosides) and nucleotides.
Overall, it is expected that the plant cytokinin diversity could be constituted out of up to one hundred
4

ACCEPTED MANUSCRIPT
derivatives. Significant and different activities have been found only in the case of cytokinin nucleosides,
which manifest antitumor, antiviral, antiprotozoal, cytokinin, blood pressure reducing, anti-inflammatory,
antipsychotic activity, influence platelet aggregation and some others [14, 27, 28, 29, 30]. One of the best
6
studied derivatives of this group is N -benzyladenosine 23, which was synthesized in the late 60ies [21],
and which only quite recently was found and characterized in plants [31].
6
Taking into consideration the earlier findings [12] and our recent observations [14], we selected N -
benzyladenosine (23) as a promising compound with anti-enterovirus activity for further optimization.
The structure of 23 has a lot of different functionalities and may be modified in several ways as marked
with the arrows on Fig. 1. In the present investigation, we have mainly focused on the size and nature of
the linker between the purine nucleus and the phenyl ring, as well as on the necessity of the D-ribose
residue. The evaluation of the antiviral activity of the compounds in an enterovirus EV71 cell-based assay
revealed that the selectivity index can be significantly improved by variation of the linker structure.
6
Fig. 1. Possible modifications of N -benzyladenosine (23).
6
N -Benzyladenosine 23 was found to be rather cytotoxic (Table 1), so, one angle to obtain compounds
with an improved selectivity index was to try to reduce the cytotoxic effect.
Table 1
Antiviral activity of adenosine derivatives on the replication of EV71 strain BrCr in RD cells.
a b
с
Compound name
Substituent (R)
CC ± SD
EC ± SD
SI
5
0
50
6
1
2
9
3
N -phenyladenosine
372 ± 16
4 ± 1
>291
ND
14
6
N -benzyladenosine
0.28 ± 0.05
5

ACCEPTED MANUSCRIPT
6
4
5
N -(2-phenylethyl)-adenosine
221 ± 32
1.8 ± 0.6
1.8 ± 0.6
123
105
6
N -(3-phenylpropan-1-yl)-
189 ± 27
426 ± 7
adenosine
6
N -(trans-3-phenyl-2-propen-1-yl)-
6
0.66 ± 0.12
645
adenosine
6
N -(3-phenyl-2-propin-1-yl)-
7
8
72 ± 4
0.58 ± 0.13
4.3 ± 0.2
124
25
adenosine
6
N -(2-phenoxyethyl)-adenosine
107 ± 40
6
2
9
4
N -(Benzyloxymethyl)adenosine
525 ± 36
>313
3.0 ± 0.3
>250
175
ND
6
N -(4-phenylbutane-1-yl)-adenosine
6
N -(3-phenoxypropan-1-yl)-
adenosine
1
2
0
5
>311
>337
>311
ND
>28
6
N -benzoyladenosine
12 ± 2
6
2
2
6
7
N -furfuryladenosine
7.8 ± 3.4
136 ± 8
1.4 ± 0.3
>136
5.6
6
N -(1-Methyl-1,2,3-triazol-4-yl-
ND
methyl)adenosine
6
N -[1-(2-hydroxyethyl)-1,2,3-
2
8
317 ± 30
>317
ND
triazol-4-yl-methyl]adenosine
6
N -(1-benzyl-1,2,3-triazol-4-yl-
2
3
9
0
>285
>228
>116
ND
ND
methyl)adenosine
6
N -(1-benzyloxymethyl-1,2,3-
116 ± 29
triazol-4-yl-methyl)adenosine
6
3
3
1
2
N -Isopenthenyladenosine
5.8 ± 0.5
>434
1.0 ± 0.2
4.9 ± 1.1
5.8
6
N -(5-hexene-2-yne-1-yl)adenosine
>88
a
All values are in ꢀM and are based on at least three independent dose-response curves.
On rhabdomyosarcoma (RD) cells
b
c
Selectivity Index (SI); SI=CC /EC
50
5
0
ND = Not Determined
We have previously shown that a hydrophobic residue is essential for antiviral activity against enterovirus
6
7
1 [14]. Therefore, we prepared different N -benzyladenosine analogs with different properties of the
linker between the purine and the phenyl group, changing from zero atoms (19) up to four (9 and 10) and
with different character, containing not only polymethylene and oxamethylene chain, but also double and
triple bonds. Our SAR study clearly shows that the optimal size of the linker appears to be limited to 2 or
6
3
atoms (compounds 4-7). Compound 8, having a more polar oxamethylene linker, was less active. N -
6

ACCEPTED MANUSCRIPT
Phenyladenosine (19), without linker, and 9 (4-atoms linker) were practically inactive, whereas 10 has
some activity.
6
Other cytokinin ribosides, 31 and 26, were less active than N -benzyladenosine 23. It is quite interesting
6
to observe that N -benzoyladenosine (25) possess anti-enterovirus 71 activity without a pronounced
adverse effect on the host cells. Compounds with triazole heterocycle (22-30) were practically inactive.
Inosine derivatives 20-21 (Table 2) were found to be much less effective than the corresponding
adenosines. Their antiviral activity decreases with an increase of the linker length.
Table 2
Antiviral effect of inosine derivatives on the replication of EV71 strain BrCr in RD cells.
a b
c
Compound name
Substituent (R)
CC ± SD
EC ± SD
SI
5
0
50
6
2
2
0
1
O -benzylinosine
191 ± 39
8.5 ± 4.1
5.1 ± 1.9
>8.5
37
6
O -(2-phenylethyl)-inosine
ND
6
O -(3-phenylpropan-1-yl)-
inosine
All values are in ꢀM and are based on at least three independent dose-response curves.
2
2
182 ± 46
>182
ND
a
b
c
On rhabdomyosarcoma (RD) cells
Selectivity Index (SI); SI=CC /EC
50
5
0
ND = Not Determined
The modulation of the length and nature of the linker between purine and phenyl group can definitely
improve the selectivity of the start compound 23 (supporting info, figures 1-3).
2
′-Deoxyadenosine derivatives 11-14 (Table 3) do not have any apparent antiviral activity or cytotoxicity.
In the 5′-deoxynucleoside series 15-16 (Table 4), the antiviral activity was substantially decreased
compared to the natural cytokinin nucleosides 23 and 31.
Table 3
Antiviral effect of 2′-deoxyadenosine derivatives on the replication of EV71 strain BrCr in RD cells.
7

ACCEPTED MANUSCRIPT
a b
c
Compound name
Substituent (R)
CC ± SD
EC ± SD
SI
5
0
50
6
N -benzyl-2′-
1
1
1
1
1
2
3
4
>366
>352
>271
>337
>146
>281
>271
>135
ND
ND
ND
ND
deoxyadenosine
6
N -(2-phenylethyl)- 2′-
deoxyadenosine
6
N -(3-phenylpropan-1-yl)-
2
′-deoxyadenosine
6
N -(2-phenoxyethyl)-2′-
deoxyadenosine
a
b
c
All values are in ꢀM and are based on at least three independent dose-response curves.
On rhabdomyosarcoma (RD) cells
Selectivity Index (SI); SI=CC /EC
50
5
0
ND = Not Determined
Table 4
Antiviral effect of 5′-deoxyadenosine derivatives on the replication of EV71 strain BrCr in RD cells.
a b
c
Compound name
Substituent (R)
CC ± SD
EC ± SD
SI
5
0
50
6
1
1
5
6
N -isopentenyl-5′-deoxyadenosine
182 ± 79
>366
13 ± 2
14
6
N -benzyl-5′-deoxyadenosine
6.7 ± 1.6
>54
6
N -(2-phenylethyl)- 5′-
1
7
463 ± 29
>281
ND
deoxyadenosine
a
b
c
All values are in ꢀM and are based on at least three independent dose-response curves.
On rhabdomyosarcoma (RD) cells
Selectivity Index (SI); SI=CC /EC
50
5
0
ND = Not Determined
6
The acyclic derivative 33 (Table 5) without intact ribose was not active compared to the related N -
benzoyladenosine (25).
Table 5
Antiviral effect of acyclic adenosine derivative on the replication of EV71 strain BrCr in RD cells.
a b
c
Compound name
Structure
CC ± SD
EC ± SD
SI
5
0
50
8

ACCEPTED MANUSCRIPT
N-(9-(1-[(1,3-
dihydroxypropan-2-yl)oxy]-2-
hydroxyethyl)-9H-purin-6-
yl)benzamide
3
3
>335
>335
ND
a
b
c
All values are in ꢀM and are based on at least three independent dose-response curves.
On rhabdomyosarcoma (RD) cells
Selectivity Index (SI); SI=CC /EC
50
5
0
ND = Not Determined
6
In the previous paper [14], we have shown that natural N -isopenthenyladenine 34 (Table 6) does not
6
protect the cells from virus-induced cell death. In contrast to this observation, N -benzyladenine (35) has
antiviral activity comparable to that of the start nucleoside 23.
Table 6
Antiviral effect of cytokinins on the replication of EV71 strain BrCr in RD cells.
a b
c
Compound name
Substituent (R)
CC ± SD
EC ± SD
SI
5
0
50
6
3
4
5
N -isopentenyladenine
325 ± 18
547 ± 14
>325
ND
6
3
N -benzyladenine
0.42 ± 0.05
1302
a
All values are in ꢀM and are based on at least three independent dose-response curves.
On rhabdomyosarcoma (RD) cells
b
c
Selectivity Index (SI); SI=CC /EC
50
5
0
ND = Not Determined
Biological activity of cytokinin nucleosides may be associated not only with the compounds as described
in this study, but also with phosphorylated metabolites that may become available following intracellular
metabolization of the parent compound. It was shown earlier that cytokinins may be converted to their
corresponding nucleoside 5′-monophosphate [32].
It was also shown [33] that 6-furyl- or 6-thienyl-7-deazapurines ribosides induced very high cytotoxicity
on several tumor cell lines and that they required 5′-triphosphorylation to be able to act by inhibition of
RNA synthesis.
9

ACCEPTED MANUSCRIPT
Our data are in line with these observations: the antiviral activity is absent for 2′-deoxyadenosine
derivatives 11-14 and the acyclic nucleoside 33. 5′-Deoxynucleosides 15-16, which do not have 5′-
hydroxyl group and could not be converted to the 5′-monophosphates, have much lower antiviral activity
than the related ribonucleosides. But the remaining activity in 15-16 points out that 5′-triphosphorylation
and inhibition of RNA synthesis cannot be the only mechanism of antiviral activity of cytokinin
6
nucleosides. The antiviral activity of N -benzyladenine 35 is in line with this conclusion.
Finally, the antiviral activity of compound 4 and 6, being the compounds with the highest selectivity
index of the entire series, was evaluated on the replication of a representative panel of clinical isolates of
EV71 (Table 7). The replication of the virus isolates that belong to genogroups B2 and B5 appears to be
2
-fold less sensitive to the antiviral effect of both compounds compared to that of genogroup C4. In turn,
both compounds more potently inhibited the replication of the virus isolate that belongs to genogroup C2.
Along the line, compound 6 consistently shows up as the most potent and selective (highest selectivity
index) inhibitor of this series.
Table 7
Antiviral effect of compounds 4 and 6 against a selection of EV71 clinical isolates.
a
EC ± SD
5
0
Genogroup
Strain
Genbank
4
6
B2
B5
C2
11316
AB575927
GQ231942
GQ231936
―
8.2 ± 2.0
5.6 ± 1.1
8.3 ± 1.0
1.9 ± 0.2
2.7 ± 1.0
3.7 ± 0.2
2.2 ± 0.1
2.0 ± 0.3
2.5 ± 0.4
0.38 ± 0.03
1.0 ± 0.4
1.16 ± 0.01
TW/96016/08
TW/70902/08
H08300 461 #812
TW/1956/05
TW/2429/04
GQ231926
GQ231927
C4
a
All values are in ꢀM and are based on at least three independent dose-response curves.
3
. Conclusions
6
We demonstrated that the antiviral activity of natural N -benzyladenosine derivatives is greatly dependent
on the size of the linker and that a linker with a length of 2-3 atoms has the most pronounced antiviral
activity. Furthermore, the nature of the linker is also of great importance, as compounds with double and
triple bonds have better activity than those with polymethylene and oxamethylene chains. As a result of
the optimization, the selectivity index could be improved with a factor of 50 (transition of 23 to 6), an
increase that is mainly to be attributed to a reduction in the adverse effect of this compound on the host
cells (increased CC ).
5
0
1
0

ACCEPTED MANUSCRIPT
4
. Experimental section
4
.1. General
The solvents and materials were reagent grade and were used without additional purification. Column
chromatography was performed on silica gel (Kieselgel 60 Merck, 0.063-0.200 mm). TLC was performed
on Alugram SIL G/UV254 (Macherey-Nagel) with UV visualization. Melting points were determined
1
13
with Electrothermal apparatus and are uncorrected. H and C (with complete proton decoupling) NMR
1
spectra were recorded on Bruker AMX 400 NMR instrument at 300K. H-NMR-spectra were recorded at
1
3
4
00 MHz and C-NMR-spectra at 100 MHz. Chemical shifts in ppm were measured relative to the
1
13
1
residual solvent signals as internal standards (CDCl , H: 7.26 ppm, C: 77.1 ppm; DMSO-d , H: 2.50
3
6
13
ppm, C: 39.5 ppm) [34]. Spin-spin coupling constants (J) are given in Hz. Double resonance technique
was applied for assigning the resonances. High resolution mass spectra (HRMS) were measured on a
Bruker micrOTOF II instrument using electrospray ionization (ESI) [35]. The measurements were done in
a positive ion mode (interface capillary voltage – 4500 V) or in a negative ion mode (3200 V); mass range
from m/z 50 to m/z 3000 Da; external or internal calibration was done with Electrospray Calibrant
Solution (Fluka). A syringe injection was used for solutions in acetonitrile, methanol, or water (flow rate
3
µL/min). Nitrogen was applied as a dry gas; interface temperature was set at 180°C.
The following compounds were prepared according to the methods reported elsewhere:
6
6
6
N -Acetyl-2′,3′,5′-tri-O-acetyladenosine (1), N -benzyladenosine (23), N -furfuryladenosine (26) were
6
6
6
prepared according to ref. [20]; N -(benzyloxymethyl)adenosine (24), O -benzylinosine (20), N -(1-
6
Methyl-1,2,3-triazol-4-yl-methyl)adenosine
(27),
N -[1-(2-hydroxyethyl)-1,2,3-triazol-4-yl-
6
6
methyl]adenosine (28), N -(1-benzyl-1,2,3-triazol-4-yl-methyl)adenosine (29), N -(1-benzyloxymethyl-
6
1
,2,3-triazol-4-yl-methyl)adenosine (30) were prepared according to ref. [27]; N -benzoyladenosine (25)
was prepared according to ref. [36]; N-(9-(1-[(1,3-dihydroxypropan-2-yl)oxy]-2-hydroxyethyl)-9H-purin-
6
4
4
-yl)benzamide (33) was prepared according to ref. [37]
.2. Adenosine derivatives
6
.2.1. N -phenyl-2′,3′,5′-tri-O-acetyl-adenosine
A mixture of 18 [24, 38] (193 mg, 0.467 mmol) and aniline hydrochloride (70 mg, 0.540 mmol) was
solved in n-BuOH (5 ml) and then DIPEA (0.185 ml, 1.08 mmol) was added in one portion. The solution
o
was stirred at 80 C. The reaction was monitored by TLC (ethyl acetate:hexane – 2:1). After 14 h, the
reaction mixture was evaporated in vacuum and the residue was diluted with methylene chloride (30 ml)
1
1

ACCEPTED MANUSCRIPT
and washed with water (2×15 ml). The organic layer was separated, dried over anhydrous Na SO and
2
4
evaporated in vacuum. The residue was purified by column chromatography on silica gel. The product
6
was eluted with CH Cl -EtOH (97:3) mixture. Yield of N -phenyl-2′,3′5′-tri-O-acetyl-adenosine was 219
2
2
1
mg (93%) as foam. R 0.42 (ethyl acetate-hexane, 2:1 v/v). H NMR (400 MHz, CDCl ): δ = 2.09 (s, 3H,
f
3
Ac), 2.13 (s, 3H, Ac), 2.15 (s, 3H, Ac), 4.37-4.49 (m, 3H, H4', H5'a, H5'b), 5.64 (dd, 1H, J3′,2′ = 5.3 Hz,
J3′,4′ = 4.5 Hz, H3'), 5.91 (dd, 1H, J2′,1′ = 5.2 Hz, J2′,3′ = 5.3 Hz, H2'), 6.22 (d, 1H, J1′,2′ = 5.2 Hz, H1'), 7.15-
7
22 (m, 1H, Hp-Ph), 7.37-7.44 (m, 2H, Hm-Ph), 7.72-7.78 (m, 2H, Ho-Ph), 8.05 (s, 1H, H8), 8.51 (s, 1H,
H2).
6
4
.2.2. N -phenyladenosine (19)
6
N -phenyl-2′,3′,5′-tri-O-acetyl-adenosine (116 mg, 0.247 mmol) was treated with 7M NH in MeOH
3
solution (3 ml). After 24 h, the mixture was evaporated in vacuum. The product was recrystallized from
MeOH. The precipitant was filtered, washed with cold MeOH and dried over P O . Yield of 19 was 69
2
5
o
1
mg (81%) as a white powder. R 0.22 (CH Cl -EtOH, 10:1 v/v). mp 199-200 C. H NMR (400 MHz,
f
2
2
DMSO-d ): δ = 3.58 (ddd, 1H, J5′b,4′ = 3.5 Hz, J5′b,OH = 6.7 Hz, J5′b,5′a = -12.1 Hz, H5'b), 3.70 (ddd, 1H,
6
J5
′a,4′
= 3.5 Hz, J5′a,OH = 4.8 Hz, J5′a,5′b = -12.1 Hz, H5'a), 3.98 (ddd, 1H, J4′,3′ = 3.5 Hz, J4′,5′a = 3.5 Hz, J4′,5′b
=
3.5 Hz, H4'), 4.19 (dd, 1H, J3′,2′ = 4.8 Hz, J3′,4′ = 3.5 Hz, H3'), 4.65 (dd, 1H, J2′,1′ = 6.0 Hz, J2′,3′ = 4.8 Hz,
H2'), 5.17 (d, 1H, J3′,OH = 4.8 Hz, 3'OH), 5.25 (dd, 1H, J5a′,OH = 4.8 Hz, J5b′,OH = 6.7 Hz, 5'OH), 5.45 (d,
H, J2′,OH = 6.2 Hz, 2'OH), 5.97 (d, 1H, J1′,2′ = 6.0 Hz, H1'), 7.02-708 (m, 1H, Hp-Ph), 7.30-7.36 (m, 2H,
1
1
3
Hm-Ph), 7.91-7.97 (m, 2H, Ho-Ph), 8.40 (s, 1H, H8), 8.53 (s, 1H, H2), 9.89 (s, 1H, NH). C NMR (100
MHz, DMSO-d ): δ = 61.56 (C5'), 70.56 (C3'), 73.60 (C2'), 85.86 (C4'), 87.86 (C1'), 120.36 (C5), 120.90
6
(Ph), 122.74 (Ph), 128.40 (Ph), 139.52 (Ph), 140.71 (C8), 149.33 (C4), 151.93 (C2), 152.17 (C6). HRMS:
+
+
m/z [M+Na] calculated C H N O Na 366.1173, found 366.1167.
16
17
5
4
6
4
.2.3. N -(2-phenylethyl)-adenosine (4)
To a solution of 1 (435 mg, 1 mmol), Ph P (393 mg, 1.5 mmol) and 2-phenylethanol (0.180 ml, 1.5
3
mmol) in THF (5 ml) DEAD (0.24 ml, 1.5 mmol) was added in one portion and the solution was kept at
ambient temperature for 20 h. The reaction was monitored by TLC (silica gel, CH Cl -EtOH, 97:3). Then,
2
2
the reaction mixture was evaporated and the residue was applied to column chromatography (silica gel,
CH Cl -EtOH = 97:3). Partially purified compound was dissolved in 4M PrNH in MeOH solution (50
2
2
2
mmol) and was left for 24 h, after which the mixture was evaporated and the residue was applied to
column chromatography. The column was washed with methylene chloride:ethanol – 95:5 and then eluted
with methylene chloride:ethanol – 90:10 to give 4 as a white powder. Yield 180 mg (65%). R 0.28
f
1
2

ACCEPTED MANUSCRIPT
o
1
(
CH Cl -EtOH, 9:1). mp 169-171 C. H NMR (400 MHz, DMSO-d ): δ = 2.93 (t, 2H, JCH2-CH2= 7.5 Hz,
2
2
6
СH Ph), 3.55 (ddd, 1H, J
= -12.0 Hz, J5′b,4′ = 3.4 Hz, J5′b,OH = 7.0 Hz, H5′b), 3.63-3.81 (m, 3H,
2
5′b,5′a
NHCH2, H5′a), 3.97 (ddd, 1H, J4′,5′b = 3.0, J4′,5′a = 3.4, J4′,3′ = 3.2, H4′), 4.15 (ddd, 1H, J3′,4′ = 3.2 Hz, J3′,2′
5.9 Hz, J3′,OH = 4.6 Hz, H3′), 4.60 (dd, 1H, J2′,3′ = 5.9 Hz, J2′,1′ = 6.1 Hz, H2′), 5.15 (d, 1H, JOH-3′ = 4.6
Hz, 3′OH), 5.36 (dd, 1H, JOH-5′b = 7.0 Hz, JOH-5′a = 4.6 Hz, 5′OH), 5.41 (d, 1H, JOH-2′ = 6.1 Hz, 2′OH),
=
5
1
7
1
3
.89 (d, 1H, J1′,2′ = 6.1 Hz, H1′),7.16-7.33 (m, 5H, Ph), 7.88 (br s, 1H, NH), 8.23 (br s, 1H, H2), 8.34 (s,
1
3
H, H8). C NMR (100 MHz, DMSO-d ): δ = 34.39 (CH Ph), 41.23 (NCH ), 61.66 (C5′), 70.63 (C3′),
6
2
2
3.49 (C2′), 85.88 (C4′), 87.94 (C1′) 119.74 (C5), 126.00 (Ph), 128.27 (Ph), 128.63 (Ph), 139.48 (Ph),
+
+
39.70 (C8), 148.27 (C4), 152.35 (C2), 154.61 (C6). HRMS: m/z [M+H] calculated C H N O
4
1
8
22
5
+
+
72.1666, found 372.1657; m/z [M+Na] calculated C H N O Na 394.1486, found 394.1474.
18
21
5
4
6
4
.2.4. N -(3-phenylpropan-1-yl)-adenosine (5)
Following the procedure for preparation of 4, condensation of 1 (300 mg, 0.68 mmol) with 3-
phenylpropan-1-ol (0.139 ml, 1.02 mmol) in the presence of Ph P (269 mg, 1.02 mmol) and DEAD (0.16
3
ml, 1.02 mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (25 mmol)
2
o
1
gave 5 as a white crystals. Yield 208 mg (79%). R 0.11 (CH Cl -EtOH, 97:3), mp 126-129 C. H NMR
f
2
2
(
400 MHz, DMSO-d ): δ = 1.91 (p, 2H, J
= 7,5 Hz, CH CH CH Ph), 2.65 (t, 2H, J
= 7,5 Hz,
6
CH2CH2
2
2
2
CH2CH2
CH Ph), 3.46-3.60 (m, 3H, H5'b, NHCH ), 3.67 (ddd, 1H, J
= 3.4 Hz, J5′a,OH = 4.0 Hz, J5′a,5′b = -12.1
2
2
5′a,4′
Hz, H5'a), 3.96 (ddd, 1H, J4′,3′ = 3.4 Hz, J4′,5′a = 3.4 Hz, J4′,5′b = 3.4 Hz, H4'), 4.15 (ddd, 1H, J3′,2′ = 4.8 Hz,
J3′,4′ = 3.4 Hz, J3′,OH = 4.6 Hz, H3'), 4.61 (dd, 1H, J2′,1′ = 6.1 Hz, J2′,3′ = 4.8 Hz, H2'), 5.14 (d, 1H, J3′,OH
=
4
7
.6 Hz, 3'OH), 5.35-5.42 (m, 2H, 2'OH, 5'OH), 5.88 (d, 1H, J1′,2′ = 6.1 Hz, H1'), 7.13-7.31 (m, 5H, Ph),
1
3
.91 (br s, 1H, NH), 8.20 (s, 1H, H8), 8.34 (s, 1H, H2). C NMR (100 MHz, DMSO-d ): δ = 30.77
6
(
CH ), 32.60 (CH2, CH2), 61.67 (C5'), 70.64 (C3'), 73.47 (C2'), 83.89 (C4'), 87.96 (C1'), 119.72 (C5),
2
1
25.64 (Ph), 128.22 (Ph), 139.60 (C8), 141.78 (Ph), 148.38 (C4), 152.31 (C2), 154.72 (C6). HRMS: m/z
+
+
+
+
[
M+H] calculated C H N O 386.1823, found 386.1814; m/z [M+Na] calculated C H N O Na
19 24 5 4 19 23 5 4
4
08.1642, found 408.1634.
6
4
.2.5. N -(trans-3-phenyl-2-propen-1-yl)-adenosine (6)
Following the procedure for preparation of 4, condensation of 1 (435 mg, 1 mmol) with trans-3-phenyl-2-
propen-1-ol (0.193 ml, 1.5 mmol) in the presence of Ph P (393 mg, 1.5 mmol) and DEAD (0.24 ml, 1.5
3
mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (50 mmol) gave 6 as a
2
o
1
white powder. Yield 238 mg (62%). R 0.08 (CH Cl -EtOH, 97:3). mp 129-132 C. H NMR (400 MHz,
f
2
2
DMSO-d ): δ = 3.56 (ddd, 1H, J5′b,4′ = 3.4 Hz, J5′b,5′a = -12.1 Hz, J5′b,OH = 6.9 Hz, H5'b), 3.68 (ddd, 1H,
6
1
3

ACCEPTED MANUSCRIPT
J5
′a,4′
= 3.4 Hz, J5′a,5′b = -12.1 Hz, J5′a,OH = 4.7 Hz, H5'a), 3.98 (ddd, 1H, J4′,3′ = 6.9 Hz, J4′,5′a = 3.4 Hz, J4′,5′b
=
3.4 Hz, H4'), 4.20 (dd, 2H, JCH2,CH=CH = 5.5 Hz (trans), JCH2,CH=CH = 2.0 Hz (cis), NHCH ), 4.62 (dd,
2
1
5
1
H, J2′,1′ = 5.9 Hz, J2′,3′ = 4.8 Hz, H2'), 5.34 (dd, 1H, J3′,2′ = 4.8 Hz, J3′,4′ = 7.1 Hz, H3'), 5.90 (d, 1H, J1′,2′
=
=
.9 Hz, H1'), 6.40 (dt, 1H, JCH=CH = 16 Hz, JCH2,CH=CH = 5.5 Hz (trans), CH CH=),6.55 (dt, 1H, J
2
CH=CH
6 Hz (trans), JCH2,CH=CH = 1.5 Hz (cis), =CHPh), 7.21 (tt, 1H, HpPh), 7.30 (dd, 2H, JHm,Ho, = 8.5 Hz,
JHm,Hp, = 7.4 Hz, HmPh), 7.39 (dd, 2H, JHo,Hm, = 8.5 Hz, JHo,Hp, = 1.2 Hz, HoPh), 8.08 (br s, 1H, NH), 8.23
1
3
(
(
(
s, 1H, H8), 8.36 (s, 1H, H2). C NMR (100 MHz, DMSO-d ): δ = 41.49 (NHCH ), 61.62 (C5'), 70.59
6 2
C3'), 73.47 (C2'), 85.85 (C4'), 87.91 (C1'), 119.81 (C5), 126.06 (Ph), 127.26 (Ph), 128.53 (Ph), 129.94
+
=CPh), 136.62 (NHCH C), 139.78 (C8), 148.38 (C4), 154.42 (C6). HRMS: m/z [M+H] calculated
2
+
+
+
C H N O 384.1666, found 384.1661; m/z [M+Na] calculated C H N O Na 406.1486, found
19
22
5
4
19 21
5
4
4
06.1479.
6
4
.2.6. N -(3-phenyl-2-propin-1-yl)-adenosine (7)
Following the procedure for preparation of 4, condensation of 1 (435 mg, 1 mmol) with 3-phenyl-2-
propin-1-ol (0.187 ml, 1.5 mmol) in the presence of Ph P (393 mg, 1.5 mmol) and DEAD (0.24 ml, 1.5
3
mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (50 mmol) gave 7 as a
2
o
1
white powder. Yield 209 mg (55%). R 0.04 (CH Cl -EtOH, 97:3). mp 128-130 C. H NMR (400 MHz,
f
2
2
DMSO-d ): δ = 3.56 (ddd, 1H, J5′b,4′ = 3.2 Hz, J5′b,5′a = -12.1 Hz, J5′b,OH = 6.8 Hz, H5'b), 3.69 (ddd, 1H,
6
J5
′a,4′
= 3.5 Hz, J5′a,5′b = -12.1 Hz, J5′a,OH = 5.3 Hz, H5'a), 3.97 (ddd, 1H, J4′,3′ = 7.2 Hz, J4′,5′a = 3.5 Hz, J4′,5′b
=
3.2 Hz, H4'), 4.16 (dd, 1H, J3′,2′ = 4.7 Hz, J3′,4′ = 7.2 Hz, H3'), 4.56 (br s, 2H, NHCH ), 4.61 (td, 1H, J2′,1′
2
=
6.1 Hz, J2′,OH′ = 6.1 Hz, J2′,3′ = 4.7 Hz, H2'), 5.29 (dd, 1H, J5′a,OH = 5.3 Hz, J5′b,OH = 6.8 Hz, 5'OH), 5.91
1
3
(
(
(
(
d, 1H, J1′,2′ = 6.0 Hz, H1'), 7.42-7.31 (m, 5H, Ph), 8.30 (br s, 2H, H8, NH), 8.40 (s, 1H, H2). C NMR
100 MHz, DMSO-d ): δ = 29.92 (NHCH ), 61.62 (C5'), 70.60 (C3'), 73.55 (C2'), 81.13 (≡C–),85.87
6
2
C4'), 87.76 (–C≡),87.93 (C1'), 119.97 (C5), 122.40 (Ph), 128.43 (Ph), 128.64 (Ph), 131.34 (Ph), 140.22
+
+
C8), 148.76 (C4), 152.29 (C2), 153.94 (C6). HRMS: m/z [M+H] calculated C H N O 382.1510,
1
9
20
5
4
+
+
found 382.1501; m/z [M+Na] calculated C H N O Na 404.1329, found 404.1321.
1
9
19
5
4
6
4
.2.7. N -(2-phenoxyethyl)-adenosine (8)
Following the procedure for preparation of 4, condensation of 1 (300 mg, 0.68 mmol) with 2-
phenoxyethanol (0.129 ml, 1.02 mmol) in the presence of Ph P (269 mg, 1.02 mmol) and DEAD (0.16
3
ml, 1.02 mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (25 mmol)
2
1
gave 8 as a white foam. Yield 153 mg (59%). R 0.04 (CH Cl -EtOH, 97:3). H NMR (400 MHz, DMSO-
f
2
2
d6): δ = 3.56 (ddd, 1H, J5′b,4′ = 4.0 Hz, J5′b,OH = 4.8 Hz, J5′b,5′a = -12.1 Hz, H5'b), 3.67 (ddd, 1H, J5′a,4′ = 3.5
1
4

ACCEPTED MANUSCRIPT
Hz, J5′a,OH = 4.8 Hz, J5′a,5′b = -12.1 Hz, H5'a), 3.88 (br s, 2H, NHCH ), 3.97 (ddd, 1H, J = 3.5 Hz, J4′,5′a
=
2
4′,3′
3
.5 Hz, J4′,5′b = 4.0 Hz, H4'), 4.25-4.10 (m, 3H, H3', CH OPh), 4.61 (ddd, 1H, J = 6.1 Hz, J2′,3′ = 4.7 Hz,
2 2′,1′
J2′,OH = 6.2 Hz, H2'), 5.14 (d, 1H, J3′,OH = 4.7 Hz, 3'OH), 5.33 (t, 1H, J5′,OH = 4.8 Hz, 5'OH), 5.39 (d, 1H,
J2′,OH = 6.2 Hz, 2'OH), 5.90 (d, 1H, J1′,2′ = 6.1 Hz, H1'), 7.00-6.87 (m, 3H, HpPh, HmPh), 7.27 (t, 2H,
1
3
JHo,Hm, = 8.0 Hz, HoPh), 7.94 (br s, 1H, NH), 8.25 (s, 1H, H8), 8.37 (s, 1H, H2). C NMR (100 MHz,
DMSO-d ): δ = 61.60 (C5'), 65.66 (NHCH ), 70.58 (C3'), 73.40 (CH OPh), 73.51 (C2'), 85.84 (C4'),
6
2
2
8
1
7.92 (C1'), 114.44 (Ph), 119.74 (C5), 120.56 (Ph), 129.45 (Ph), 139.67 (C8), 148.44 (C4), 152.28 (C2),
+
+
5
54.67 (C6), 158.39 (OPh). HRMS: m/z [M+H] calculated C H N O 388.1615, found 388.1612; m/z
1
8
22
5
+
+
[
M+Na] calculated C H N O Na 410.1435, found 410.1431.
18 21 5 5
6
4
.2.8. N -(4-phenylbutane-1-yl)-adenosine (9)
Following the procedure for preparation of 4, condensation of 1 (250 mg, 0.57 mmol) with 4-
phenylbutane-1-ol (0.174 ml, 1.14 mmol) in the presence of Ph P (224 mg, 0.85 mmol) and DEAD (0.133
3
ml, 0.85 mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (25 mmol)
2
o
1
gave 9 as a white powder. Yield 242 mg (60%). R 0.18 (CH Cl -EtOH, 97:3). mp 133-134 C. H NMR
f
2
2
(
400 MHz, DMSO-d ): δ = 1.57-1.65 (m, 4H, CH CH CH CH Ph), 2.60 (t, 2H, J
= 6,8 Hz,
6
2
2
2
2
CH2CH2
CH Ph), 3.45-3.60 (m, 3H, H5'b, NHCH ), 3.68 (ddd, 1H, J
= 3.4 Hz, J5′a,OH = 4.2 Hz, J5′a,5′b = -12.1
2
2
5′a,4′
Hz, H5'a), 3.97 (ddd, 1H, J4′,3′ = 3.4 Hz, J4′,5′a = 3.4 Hz, J4′,5′b = 3.4 Hz, H4'), 4.15 (ddd, 1H, J3′,2′ = 5.2 Hz,
J3′,4′ = 3.4 Hz, J3′,OH = 4.6 Hz, H3'), 4.61 (dd, 1H, J2′,1′ = 6.2 Hz, J2′,3′ = 5.2 Hz, H2'), 5.14 (d, 1H, J3′,OH
=
4
7
.6 Hz, 3'OH), 5.35-5.41 (m, 2H, 2'OH, 5'OH), 5.88 (d, 1H, J1′,2′ = 6.2 Hz, H1'), 7.12-7.30 (m, 5H, Ph),
1
3
.87 (br s, 1H, NH), 8.19 (s, 1H, H8), 8.32 (s, 1H, H2). C NMR (100 MHz, DMSO-d ): δ = 28.37
6
(
(
(
CH ), 28.72 (CH ), 34.84 (CH ), 61.66 (C5'), 70.63 (C3'), 73.45 (C2'), 85.87 (C4'), 87.95 (C1'), 119.66
2 2 2
C5), 125.55 (Ph), 128.14 (Ph), 128.24 (Ph), 139.55 (C8), 142.16 (Ph), 148.16 (C4), 152.30 (C2), 154.66
C6).
+
+
4
+
HRMS: m/z [M+H] calculated C H N O 400.1979, found 400.1976; m/z [M+Na] calculated
2
0
26
5
+
+
+
C H N O Na 422.1799, found 422.1795; m/z [M+K] calculated C H N O K 438.1538, found
20
25
5
4
20 25
5
4
4
38.1547.
6
4
.2.9. N -(3-phenoxypropan-1-yl)-ladenosine (10)
Following the procedure for preparation of 4, condensation of 1 (350 mg, 0.79 mmol) with 3-
phenoxypropan-1-ol (0.233 ml, 1.59 mmol) in the presence of Ph P (310 mg, 1.18 mmol) and DEAD
3
(
0.186 ml, 1.18 mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (40
2
o
1
mmol) gave 10 as a white powder. Yield 116 mg (29%). R 0.18 (CH Cl -EtOH, 97:3). mp 113-116 C. H
f
2
2
1
5

ACCEPTED MANUSCRIPT
NMR (400 MHz, DMSO-d ): δ = 2.07 (p, 2H, J
= 6,5 Hz, CH CH CH ), 3.55 (ddd, 1H, J
= 3.2
6
CH2CH2
2
2
2
5′b,4′
Hz, J5′b,OH = 6.9 Hz, J5′b,5′a = -12.1 Hz, H5'b), 3.68 (ddd, 1H, J5′a,4′ = 3.2 Hz, J5′a,OH = 4.5 Hz, J5′a,5′b = -12.1
Hz, H5'a), 3.62-3.73 (m, 2H, NHCH ), 3.97 (ddd, 1H, J = 3.2 Hz, J4′,5′a = 3.2 Hz, J4′,5′b = 3.2 Hz, H4'),
2
4′,3′
4
.05 (t, JCH2CH2 = 6,5 Hz, CH OPh), 4.15 (ddd, 1H, J = 4.7 Hz, J3′,4′ = 3.2 Hz, J3′,OH = 4.5 Hz, H3'), 4.61
2 3′,2′
(ddd, 1H, J2′,1′ = 5.9 Hz, J2′,3′ = 4.7 Hz, J2′,OH = 6.1 Hz, H2'), 5.13 (d, 1H, J3′,OH = 4.5 Hz, 3'OH), 5.35 (dd,
1
H, J5a′,OH = 4.5 Hz, J5b′,OH = 6.9 Hz, 5'OH), 5.38 (d, 1H, J2′,OH = 6.1 Hz, 2'OH), 5.89 (d, 1H, J1′,2′ = 5.9 Hz,
1
3
H1'), 6.88-7.31 (m, 5H, Ph), 7.92 (br s, 1H, NH), 8.20 (s, 1H, H8), 8.34 (s, 1H, H2). C NMR (100 MHz,
DMSO-d ): δ = 28.81 (CH CH CH ), 37.07 (NHCH ), 61.64 (C5'), 65.39 (CH O), 70.61 (C3'), 73.48
6
2
2
2
2
2
(C2'), 85.86 (C4'), 87.95 (C1'), 114.43 (Ph), 119.83 (C5), 120.39 (Ph), 129.38 (Ph), 139.67 (C8), 148.29
(C4), 152.30 (C2), 154.71 (C6), 158.60 (OPh).
+
+
5
+
HRMS: m/z [M+H] calculated C H N O 402.1772, found 402.1763; m/z [M+Na] calculated
1
9
24
5
+
+
+
C H N O Na 424.1591, found 424.1583; m/z [M+K] calculated C H N O K 440.1331, found
19
23
5
5
19 23
5
5
4
4
4
40.1329.
.3. 2′-Deoxyadenosine derivatives
6
.3.1. N -benzyl-2′-deoxyadenosine (11)
A mixture of 2 (194 mg, 0.5 mmol), benzyl bromide (0.089 ml, 0.75 mmol) and DBU (0.149 ml, 1 mmol)
in dry acetonitrile (5 ml) was kept at ambient temperature for 20 h. The reaction mixture was neutralized
with acetic acid to pH -6.5 and evaporated in vacuum. The residual syrup was diluted with ethyl acetate
(15 ml) and washed successively with brine (2×10 ml), 10% aqueous sodium bicarbonate (20 ml) and
water (2×10 ml). The organic layer was separated, dried over anhydrous sodium sulfate and evaporated in
vacuum. The residue was applied to column chromatography. The product was eluted with methylene
chloride:ethanol – 98:2. Purified triacetyl compound was dissolved in 4M PrNH in MeOH solution (25
2
mmol) and was left for 24 h, after which the mixture was evaporated and the residue was applied to
column chromatography. The column was washed with methylene chloride:ethanol – 95:5 and then eluted
with methylene chloride:ethanol – 90:10 to give 11 as a white foam. Yield 93 mg (55%). R 0.08
f
1
(
CH Cl -EtOH, 97:3). H NMR (400 MHz, DMSO-d ): δ = 2.25 (ddd, 1H, J
= 5.8 Hz, J2′b,3′ = 2.3 Hz,
2
2
6
2′b,1′
J2′b,2′a = -13.1 Hz, H2'b), 2.74 (ddd, 1H, J2′a,1′ = 7.8 Hz, J2′a,3′ = 5.7 Hz, J2′a,2′b = -13.1 Hz, H2'a), 3.51 (ddd,
H, J5′b,4′ = 6.2 Hz, J5′b,5′a = -12.0 Hz, J5′b,OH = 6.4 Hz, H5'b), 3.62 (ddd, 1H, J5′a,4′ = 3.9 Hz, J5′a,5′b = -12.0
Hz, J5′a,OH = 5.2 Hz, H5'a), 3.88 (ddd, 1H, J4′,3′ = 2.3 Hz, J4′,5′a = 3.9 Hz, J4′,5′b = 6.2 Hz, H4'), 4.45-4.37 (m,
1
1
3
1
H, H3'), 4.70 (br s, 2H, NHCH ), 5.20 (dd, 1H, J
= 5.2 Hz, JOH,5′b = 6.4 Hz, 5'OH), 5.3 (d, 1H,
2
OH, 5′a
'OH), 6.35 (dd, 1H, J1′,2′b = 5.9 Hz, J1′,2′a = 7.8 Hz, H1'), 7.37-7.16 (m, 5H, Ph), 8.19 (s, 1H, H8), 8.35 (s,
1
3
H, H2), 8.41 (br s, 1H, NH). C NMR (400 MHz, DMSO-d ): δ = 154.90 (C6), 152.86 (C2), 148.61
6
1
6

ACCEPTED MANUSCRIPT
(
C4), 140.10 (C8), 128.82 (Ph), 127.58 (Ph), 127.32 (Ph), 119.55 (C5), 88.28 (C1'), 84.62 (C4'), 71.34
+
(
C3'), 62.20 (C5'), 43.38 (NHCH ), 39.49 (C2' overlapping with DMSO). HRMS: m/z [M+H] calculated
2
+
+
+
C H N O 342.1561, found 342.1564; m/z [M+Na] calculated C H N O Na 364.1380, found
17
20
5
3
17 19
5
3
3
64.1377.
6
4
.3.2. N -(2-phenylethyl)-2′-deoxyadenosine (12)
Following the procedure for preparation of 4, condensation of 2 (380 mg, 1 mmol) with 2-phenylethanol
0.180 ml, 1.5 mmol) in the presence of Ph P (393 mg, 1.5 mmol) and DEAD (0.24 ml, 1.5 mmol) in
(
3
THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (50 mmol) gave 12 as a white
2
1
foam. Yield 206 mg (58%). R 0.06 (CH Cl -EtOH, 97:3). H NMR (400 MHz, DMSO-d ): δ = 2.72
f
2
2
6
(ddd, 1H, J2′b,1′ = 6.1 Hz, J2′b,3′ = 2.9 Hz, J2′b,2′a = -13.1 Hz, H2'a), 2.72 (ddd, 1H, J2′a,1′ = 7.3 Hz, J2′a,3′ = 5.6
Hz, J2′a,2′b = -13.1 Hz, H2'a), 2.92 (dd, 2H, JCH2-CH2 = 7.8 Hz, J CH2-CH2 = 7.0 Hz, CH Ph), 3.52 (ddd, 1H,
2
J5′b,4′ = 4.1 Hz, J5′b,5′a = -12.0 Hz, JOH,5′b = 6.4 Hz, H5'b), 3.62 (ddd, 1H, J5′a,4′ = 4.3 Hz, J5′a,5′b = -12.0 Hz,
JOH,5′a = 5.1 Hz, H5'a), 3.82-3.60 (m, 2H, NHCH ), 3.88 (ddd, 1H, J = 6.6 Hz, J4′,5′a = 4.3 Hz, J4′,5′b = 4.1
2
4′,3′
Hz, H4'), 4.44-4.36 (m, 1H, H3'), 5.18 (dd, 1H, JOH,5′b = 6.4 Hz, JOH,5′a = 5.1 Hz, 5'OH), 5.27 (d, 1H,
JOH,3′ = 3.9 Hz, 3'OH), 6.35 (dd, 1H, J1′,2′b = 6.2 Hz, J1′,2′a = 7.3 Hz, H1'), 7.35-7.15 (m, 5H, Ph), 7.83 (br s,
1
3
1
H, NH), 8.22 (s, 1H, H8), 8.32 (s, 1H, H2). C NMR (100 MHz, CD OD): δ = 36.68 (CH Ph), 41.58
3 2
(
C2'), 43.19 (NHCH ), 63.67 (C5'), 73.07 (C3'), 87.15 (C4'), 89.91 (C1'), 121.30 (C5), 127.31 (Ph),
2
1
29.46 (Ph), 129.88 (Ph), 140.47 (Ph), 140.93 (C8), 148.86 (C4), 153.51 (C2), 156.21 (C6). HRMS: m/z
+
+
+
+
[
M+H] calculated C H N O 356.1717, found 356.1712; m/z [M+Na] calculated C H N O Na
18 22 5 3 18 21 5 3
3
78.1537, found 378.1528.
6
4
.3.3. N -(3-phenylpropan-1-yl)-2′-deoxyadenosine (13)
Following the procedure for preparation of 4, condensation of 2 (195 mg, 0.5 mmol) with 3-
phenylpropan-1-ol (0.102 ml, 0.75 mmol) in the presence of Ph P (196 mg, 0.75 mmol) and DEAD
3
(
0.117 ml, 0.75 mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (25
2
1
mmol) gave 13 as a white foam. Yield 106 mg (57%). R 0.11 (CH Cl -EtOH, 97:3). H NMR (400 MHz,
f
2
2
DMSO-d ): δ = 1.90 (p, 2H, J
= 7,6 Hz, CH CH CH ), 2.26 (ddd, 1H, J
= 6.1 Hz, J2′b,3′ = 2.9
6
CH2CH2
2
2
2
2′b,1′
Hz, J2′b,2′a = -13.0 Hz, H2'b), 2.64 (t, 2H, JCH2CH2 = 7,6 Hz, CH Ph), 2.72 (ddd, 1H, J
= 7.8 Hz, J2′a,3′ =
2
2′a,1′
5
1
4
.6 Hz, J2′a,2′b = -13.0 Hz, H2'a), 3.58-3.45 (m, 3H, H5'b, NHCH ), 3.63 (ddd, 1H, J
= 4.6 Hz, J5′a,5′b = -
2
5′a,4′
2.0 Hz, J5′a,OH = 5.0 Hz, H5'a), 3.88 (ddd, 1H, J4′,3′ = 4.1 Hz, J4′,5′a = 4.6 Hz, J4′,5′b = 3.9 Hz, H4'), 4.45-
.38 (m, 1H, H3'), 5.19 (t, 1H, JOH,5′ab = 7.0 Hz, 5'OH), 5.26 (d, 1H, 3'OH), 6.35 (dd, 1H, J1′,2′ b = 6.1 Hz,
1
3
J1′,2′ a = 7.8 Hz, H1'), 7.30-7.10 (m, 5H, Ph), 7.85 (br s, 1H, NH), 8.18 (s, 1H, H8), 8.31 (s, 1H, H2). C
1
7

ACCEPTED MANUSCRIPT
NMR (100 MHz, DMSO-d6): δ = 32.60, 30.79 (NHCH CH CH Ph), 39.50 (C2' overlapping with
2
2
2
DMSO), 61.89 (C5'), 70.96 (C3'), 83.95 (C4'), 87.99 (C1'), 119.62 (C5), 125.63 (Ph), 128.22 (Ph), 139.21
+
(
C8), 141.79 (Ph), 148.09 (C4), 152.29 (C2), 154.68 (C6). HRMS: m/z [M+Na] calculated
+
+
+
C H N O Na 392.1693, found 392.1691; m/z [M+K] calculated C H N O K 408.1432, found
19
23
5
3
19 23
5
3
4
08.1427.
6
4
.3.4. N -(2-phenoxyethyl)-2′-deoxyadenosine (14)
Following the procedure for preparation of 4, condensation of 2 (195 mg, 0.5 mmol) with 2-
phenoxyethanol (0.093 ml, 0.75 mmol) in the presence of Ph P (196 mg, 0.75 mmol) and DEAD (0.117
3
ml, 0.75 mmol) in THF (5 ml) with subsequent deblocking in 4M PrNH in MeOH solution (25 mmol)
2
1
gave 14 as a white foam. Yield 135 mg (72%). R 0.04 (CH Cl -EtOH, 97:3). H NMR (400 MHz,
f
2
2
DMSO-d ): δ = 2.28 (ddd, 1H, J2′b,1′ = 6.4 Hz, J2′b,3′ = 2.9 Hz, J2′b,2′a = -13.1 Hz, H2'b), 2.72 (ddd, 1H, J2′a,1′
6
=
7.5 Hz, J2′a,3′ = 5.6 Hz, J2′a,2′b = -13.1 Hz, H2'a), 3.52 (ddd, 1H, J5′b,4′ = 4.2 Hz, J5′b,5′a = -12.0 Hz, JOH,5′b
.5 Hz, H5'b), 3.62 (ddd, 1H, J5′a,4′ = 4.2 Hz, J5′a,5′b = -12.0 Hz, JOH,5′a = 5.1 Hz, H5'a), 3.95-3.80 (m, 2H,
CH CH OPh), 3.88 (ddd, 1H, J = 3.4 Hz, J4′,5′a = 4.2 Hz, J4′,5′b = 4.2 Hz, H4'), 4.18 (t, 2H, JCH2CH2 = 6.2
=
6
2
2
4′,3′
Hz, CH CH OPh), 4.41 (ddd, 1H, J = 5.6 Hz, J3′,2b′ = 2.9 Hz, J3′,4′ = 3.4 Hz, H3'), 5.15 (dd, 1H, JOH,5′b
3′,2a′
2
2
=
6.5 Hz, JOH,5′a = 5.1 Hz, 5'OH), 5.26 (d, 1H, JOH,3′ = 4.0 Hz, 3'OH), 6.35 (dd, 1H, J1′,2′b = 6.4 Hz, J1′,2′a
=
7
7
.6 Hz, H1'), 7.00-6.85 (m, 3H, HoPh, HpPh), 7.28 (dd, 2H, JHm,Ho, = 8.4 Hz, JHm,Hp, = 7.2 Hz, HmPh),
1
3
.90 (br s, 1H, NH), 8.24 (s, 1H, H8), 8.35 (s, 1H, H2). C NMR (100 MHz, DMSO-d ): δ = 39.50 (C2',
6
NHCH2 overlapping with DMSO), 61.85 (C5'), 65.70 (CH O), 70.92 (C3'), 83.92 (C4'), 87.99 (C1'),
2
1
14.43 (Ph), 119.71 (C5), 120.56 (Ph), 129.45 (Ph), 139.52 (C8), 148.28 (C4), 152.27 (C2), 154.62 (C6),
+
+
+
1
58.39 (OPh). HRMS: m/z [M+H] calculated C H N O 372.1666, found 372.1659; m/z [M+Na]
18
22
5
4
+
calculated C H N O Na 394.1486, found 394.1476.
1
8
21
5
4
4
4
.4. Inosine derivatives
6
.4.1. O -(2-phenylethyl)-inosine (21)
To the mixture of 18 [36, 38] (300 mg, 0.727 mmol) and 2-phenylethanol (7.5 ml, 61.8 mmol), t-BuOK
816 mg, 7.27 mmol) was gradually added. The mixture was stirred at room temperature for 24 h. After
(
neutralization with AcOH (0.42 ml, 61.8 mmol), the solution was directly subjected to column
chromatography. Elution with a gradient of EtOH in CH Cl (1:100→1:8) gave 21 (252 mg, 93%) as a
2
2
o
1
white powder. R 0.3 (CH Cl -EtOH, 9:1). mp 154-156 C. H NMR (400 MHz, DMSO-d +D O): δ =
f
2
2
6
2
3
.15 (t, 2Н, JCH2-CH2 = 6.9 Hz, CH Ph), 3.57 (ddd, 1H, J
= -12.0 Hz, J5′b,4′ = 3.6 Hz, J5′b,OH = 5.3 Hz,
2
5′b,5′a
1
8

ACCEPTED MANUSCRIPT
H5′b), 3.68 (ddd, 1H, J5′a,5′b = -12.0 Hz, J
= 3.6 Hz, J
= 5.3 Hz, H5′a), 3.98 (ddd, 1H, J
= 3.6
5′a,4′
5′a,OH
4′,5′b
Hz, J4′,5′a = 3.6 Hz, J4′,3′ = 3.6 Hz, H4′), 4.17 (dd, 1H, J3′,4′ = 3.6 Hz, J3′,2′ = 4.4 Hz, H3′), 4.58 (dd, 1H, J2′,3′
4.4 Hz, J2′,1′ = 5.7 Hz, H2′), 4.78 (t, 2Н, JCH2-CH2= 6.9 Hz, OCH ), 5.10 (t, 1H, J = 5.3 Hz, 5′OH),
=
2
5′,OH
5
.18 (br s, 1H, 3′OH), 5.46 (br s, 1H, 2′OH), 5.98 (d, 1H, J1′,2′ = 5.7 Hz, H1′), 7.15-7.37 (m, 5H, Ph), 8.53
1
3
(
(
(
s, 1H, H2), 8.60 (s, 1H, H8). C NMR (100 MHz, DMSO-d ): δ = 34.50 (CH Ph), 61.31 (C5′), 67.04
6 2
OCH ), 70.32 (C3′), 73.77 (C2′), 85.70 (C4′), 87.80 (C1′), 121.08 (C5), 126.35 (Ph), 128.35 (Ph), 128.88
2
+
Ph), 137.96 (Ph), 142.36 (C8), 151.61 (C2), 151.89 (C4), 160.00 (C6). HRMS: m/z [M+H] calculated
+
+
+
C H N O 373.1506, found 373.1505; m/z [M+Na] calculated C H N O Na 395.1326, found
18
21
4
5
18 20
4
5
3
95.1317.
6
4
.4.2. O -(3-phenylpropan-1-yl)-inosine (22)
6
The synthesis was analogous to O -(2-phenylethyl)-inosine (21) with 3-phenyl-1-propanol (3 ml, 29
1
mmol). Yield 140 mg (90%) as a foam. R 0.11 (CH Cl -EtOH, 95:5). H NMR (400 MHz, DMSO-d ): δ
f
2
2
6
=
2.13 (ddt, 2H, JCH2-OCH2 = 6.6 Hz, JCH2-CH2Ph = 6.6 Hz, JCH2-CH2Ph= 7.9 Hz, CH CH CH ), 2.77 (dd, 2H,
2 2 2
JCH2-CH2 = 6.6 Hz, JCH2-CH2= 7.9 Hz, CH Ph), 3.58 (ddd, 1H, J
= -12.0 Hz, J5′b,4′ = 3.7 Hz, J5′b,OH = 5.6
2
5′b,5′a
Hz, H5′b), 3.68 (ddd, 1H, J5′b,5′a = -12.0 Hz, J5′a,4′ = 3.7 Hz, J5′a,OH = 5.4 Hz, H5′a), 3.98 (ddd, 1H, J4′,5′b
=
3
.7 Hz, J4′,5′a = 3.7 Hz, J4′,3′ = 3.7 Hz, H4′), 4.17 (m, 1H, H3′), 4.55 (t, 2H, JOCH2-CH2 = 6.6 Hz, OCH2),
.55-4.63 (m, 1H, H2′), 5.12 (dd, 1H, J5′b,OH = 5.6 Hz, J5′a,OH = 5.4 Hz, 5'OH), 5.20 (br s, 1H, 3'OH), 5.48
4
(d, 1H, J2′,OH = 5.0 Hz, 2'OH), 5.99 (d, 1H, J1′,2′ = 5.7 Hz, H1′), 7.16-7.32 (m, 5H, Ph), 8.52 (s, 1H, H8),
1
3
8
.62 (s, 1H, H2). C NMR (100 MHz, DMSO-d ): δ = 29.91 (CH ), 31.34 (CH ), 61.33 (C5′), 65.95
6 2 2
(
OCH ), 70.34 (C3′), 73.76 (C2′), 85.71 (C4′), 87.81 (C1′), 121.12 (C5), 125.86 (Ph), 128.27 (Ph), 128.34
2
+
(
Ph), 141.40 (Ph), 142.34 (C8), 151.61 (C2), 151.85 (C4), 160.15 (C6). HRMS: m/z [M+Na] calculated
+
+
+
C H N O Na 409.1482, found 409.1483; m/z [M+K] calculated C H N O K 425.1222, found
19
22
4
5
19 22
4
5
4
4
4
25.1226.
.5. 5′-Deoxyadenosine derivatives
.5.1. 5′-chloro-5′-deoxyadenosine (36)
To a cold suspension (0°С) of adenosine (3.0 g, 11.2 mmol) in acetonitrile (35 ml) and pyridine (22.4
mmol, 1.8 ml), thionyl chloride (4.1 ml, 56.1 mmol) was slowly added and the mixture was stirred 4h
under 0°С. The resulting solution was kept at ambient temperature overnight. The precipitate was filtered
and dissolved in H O/MeOH (5:50 ml) and 25% aqueous ammonia (4.7 ml) was added to the mixture.
2
The reaction mixture was kept at ambient temperature for 30 min and evaporated in vacuum. The residue
1
9

ACCEPTED MANUSCRIPT
was transferred to a glass filter, washed with cold water (2×20 ml) and dried in vacuum desiccator over
1
P O . Yield 1.92 g (60%) as white crystals. R 0.15 (CH Cl -EtOH, 9:1 v/v). H NMR (DMSO-d ): δ =
2
5
f
2
2
6
3
.84 (dd, 1H, J5'b-4' = 6.3 Hz, J5'ba = -11.6 Hz, H5'b), 3.94 (dd, 1H, J5'a-4' = 5.1 Hz, J5'ab = -11.6 Hz, H5'a),
.10 (ddd, 1H, J4'-5'b = 6.3 Hz, J4'-5'a = 5.1 Hz, J4'-3' = 4 Hz, H4'), 4.23 (ddd, 1H, J3'-4' = 4 Hz, J3'-OH = 5.1
4
Hz, J3'-2' = 5 Hz, H3'), 4.75 (dd, 1H, J2'-3' = 5 Hz, J2'-1' = 5.6 Hz, H2'), 5.41 (d, 1H, JOH-3' = 5.1 Hz, 3'OH),
5
.56 (d, 1H, JOH-2' = 6.03 Hz, 2'OH), 5.93 (d, 1H, J1'-2' = 5.6 Hz, H1'), 7.26 (s, 2H, NH ), 8.15 (s, 1H, H8),
2
8
.33 (s, 1H, H2).
4
.5.2. 2′,3′-Di-O-acetyl-5′-chloro-5′-deoxyadenosine (37)
To a cold solution (0°С) of 36 (1.2 g, 4.1 mmol) in 10 ml of pyridine, acetic anhydride (0.85 ml, 9.0
mmol) was added under stirring and the mixture was kept at ambient temperature for 24 h. Subsequently,
ethanol (10 ml) was added to the mixture. The resulting solution was kept at ambient temperature for 10
min and evaporated in vacuum. The residue was co-evaporated with toluene (2×40 ml) and ethanol (2×30
ml), transferred to a glass filter, washed with cold ethanol (15 ml) and dried in vacuum desiccator over
P O to give 1.21 g (80%) of 2′,3′-di-O-acetyl-5′-chloro-5′-deoxyadenosine as white powder. R 0.23
2
5
f
1
(
CH Cl -EtOH, 95:5). H NMR (DMSO-d ): δ = 2.02 (s, 3H, Ac), 2.11 (s, 3H, Ac), 3.85 (dd, 1H, J
=
2
2
6
5'b-4'
4
.21 Hz, J5'ba = -12.15 Hz, H5'b), 3.89 (dd, 1H, J5'a-4' = 4.36 Hz, J5'ab = -12.14 Hz, H5'a), 4.46 (ddd, 1H,
J4'-5'b = 4.2 Hz, J4'-5'a = 4.3 Hz, J4'-3' = 3.7 Hz, H4'), 5.61 (dd, 1H, J3'-4' = 3.7 Hz, J3'-2' = 5.6 Hz, H3'), 5.85
dd, 1H, J2'-3' = 5.6 Hz, J2'-1' = 6.07 Hz, H2'), 6.20 (d, 1H, J1'-2' = 6.07 Hz, H1'), 8.05 (s, 1H, H2), 8.28 (s,
(
1
H, H8).
6
4
.5.3. N -Acetyl-2′,3′-di-O-acetyl-5′-deoxyadenosine (3)
A mixture of 37 (400 mg, 1.08 mmol), α,α′-azobisisobutyronitrile (20 mg) and tributyltin hydride (1.46
ml, 5.4 mmol) in 20 ml of dry toluene was boiled during 2 h. The solution was evaporated in vacuum and
the residue was chromatographed on a column, containing 60 ml of silica-gel. The column was washed
with methylene chloride:ethanol – 95:5. The reaction product was eluted with methylene chloride:ethanol
–
93:7. The fractions, containing the product, were collected and evaporated in vacuo to dryness. The
residue was dissolved in pyridine (5 ml) and acetic anhydride (1 ml, 10.8 mmol) was added to the
solution under stirring. The reaction mixture was kept at 60°С for 4h. The excess of acetic anhydride was
neutralized by ethanol (20 ml). The reaction mixture was evaporated in vacuum, co-evaporated with
toluene (2×40 ml) and ethanol (2×30 ml). The residue was dissolved in ethanol (5 ml) and imidazole (70
mg, 1.03 mmol) was added to the solution. The solution was kept at ambient temperature for 4h after
which it was diluted with water (20 ml) and poured into a separating funnel. The product was extracted by
2
0

ACCEPTED MANUSCRIPT
ethyl acetate (10×10 ml). The organic layers were separated, collected, dried over anhydrous sodium
sulfate and evaporated in vacuo to dryness. Yield 310 mg (76%) as white foam. R 0.62 (methylene
f
1
chloride:ethanol – 97:3). H NMR (400 MHz, CDCl ): δ = 1.52 (d, 3H, J
= 6.4 Hz, 5'-CH ), 2.08 (s,
3
3
CH3-4'
3
1
5
H, Ac), 2.14 (s, 3H, Ac), 2.64 (s, 3H, Ac), 4.36 (qd, 1H, J4'-CH3 = 6.4 Hz, J4'-3' = 5.2 Hz, H4'), 5.40 (dd,
H, J3'-2' = 5.3 Hz, J3'-4' = 5.2 Hz, H3'), 5.96 (dd, 1H, J2'-1' = 5.0 Hz, J2'-3' = 5.3 Hz, H2'), 6.14 (d, 1H, J1'-2'
.0 Hz, H1'), 8.18 (s, 1H, H8), 8.71 (s, 1H, H2), 8.96 (br s, 1H, NH).
=
6
6
4
.5.4. N -Benzyl-N -acetyl-2′,3′-di-O-acetyl-5′-deoxyadenosine (38)
A mixture of 3 (282 mg, 0.75 mmol), benzyl bromide (0.178 ml, 1.5 mmol) and DBU (0.224 ml, 1.5
mmol) in dry acetonitrile (5 ml) was kept at ambient temperature for 3 days. The reaction mixture was
neutralized with acetic acid to pH -6.5 and evaporated in vacuum. The residual syrup was diluted with
ethyl acetate (15 ml) and washed successively with brine (2×10 ml), 10% aqueous sodium bicarbonate
(20 ml) and water (2×10 ml). The organic layer was separated, dried over anhydrous sodium sulfate and
evaporated in vacuum. The residue was chromatographed on a column containing 25 ml of silica-gel. The
product was eluted with methylene chloride:ethanol – 98:2. Yield 282 mg (81%) as white foam. R 0.49
f
1
(
CH Cl -EtOH, 98:2). H NMR (400 MHz, CDCl ): δ = 1.51 (d, 3H, J
= 6.6 Hz, 5'-CH ), 2.08 (s, 3H,
CH3-4' 3
2
2
3
Ac), 2.14 (s, 3H, Ac), 2.34 (s, 3H, Ac), 4.34 (qd, 1H, J4'-CH3 = 6.6 Hz, J4'-3' = 5.6 Hz, H4'), 5.41 (dd, 1H,
J3'-2' = 5.4 Hz, J3'-4' = 5.6 Hz, H3'), 5.54 (br s, 2H, CH Ph), 5.94 (dd, 1H, J2'-1' = 4.8 Hz, J2'-3' = 5.4 Hz,
2
H2'), 6.12 (d, 1H, J1'-2' = 4.8 Hz, H1'), 7.14 (tt, 1H, Jp-H-m-H = 8.5 Hz, Jp-H-o-H = 1.3 Hz, Ph), 7.21 (dd, 2H,
Jo-H-m-H = 7.6 Hz, Jo-H-p-H = 1.1 Hz, Ph), 7.28 (dd, 2H, Jm-H-o-H = 7.6 Hz, Jm-H-p-H = 8.5 Hz, Ph), 8.14 (s, 1H,
H8), 8.77 (s, 1H, H2).
6
4
.5.5. N -benzyl-5′-deoxyadenosine (16)
A solution of 38 (241 mg, 0.51 mmol) in 5M propylamine in methanol (5 ml) was kept for 24 h at 20°С
and then concentrated in vacuo to dryness. The residue was evaporated with ethanol (5×10 ml). The
product was precipitated from mixture methylene chloride:hexane:ethanol – 63:34:3 (3 ml). The
precipitate was filtered, washed with mixture methylene chloride:hexane:ethanol – 63:34:3 (4×2 ml) and
dried in a vacuum desiccator over P O to give 153 mg (88%) of 16 as a white powder. R 0.32 (CH Cl -
2
5
f
2
2
o
1
EtOH, 95:5). mp 198-201 C. H NMR (400 MHz, DMSO-d ): δ = 1.30 (dd, 3H, JCH3-4'= 4.77 Hz, JCH3-3'OH
6
=
1.4 Hz, CH ), 3.93-4.10 (m, 2H, H3', H4'), 4.60-4.83 (m, 3H, H2', CH Ph), 5.10 (d, 1H, J
= 4.61
3
2
2'-OH
Hz, 2'OH), 5.38 (d, 1H, J3'-OH = 5.25 Hz, 3'OH), 5.85 (dd, 1H, J1'-2' = 4.77 Hz, J1'-2'OH = 2.54 Hz, H1'),
1
3
7
.16-7.37 (m, 5H, Ph), 8.21 (s, 1H, H-8), 8.29-8.37 (m, 2H, H-2, NH). C NMR (100 MHz, DMSO-d ):
6
δ = 16.89 (C5'), 42.89 (CH Ph), 73.05 (C3'), 74.59 (C2'), 79.71 (C4'), 87.92 (C1'), 119.51 (C5), 126.54
2
2
1

ACCEPTED MANUSCRIPT
(
Ph), 127.08 (Ph), 128.14 (Ph), 139.82 (C8), 140.04 (Ph), 148.79 (C4), 152.54 (C2), 154.47 (C6). HRMS:
+
+
m/z [M+Na] calculated C H N O Na 364.1380, found 364.1374.
17
19
5
3
6
4
.5.6. N -isopentenyl-5′-deoxyadenosine (15)
6
6
The alkylation was analogous to N -Benzyl-N -acetyl-2′,3′-di-O-acetyl-5′-deoxyadenosine (38) with
isopentenyl bromide (0.232 ml, 1.06 mmol). The deacetylation analogous to 16 yielded 80 mg (47% for
o
1
two steps) of 15 as a white powder. R 0.31 (CH Cl -EtOH, 95:5). mp 117-120 C. H NMR (400 MHz,
f
2
2
DMSO-d ): δ = 1.30 (d, 3H, J
= 6.2 Hz, CH ), 1.67 (d, 3H, J
= 0.8 Hz, Me-trans), 1.70 (d, 3H,
6
CH3-CH
3
CH3-CH
JCH3-CH = 0.5 Hz, Me-cis), 3.93-4.01 (m, 2H, H3', H4'), 4.08 (br s, 2H, NHCH ), 4.65 (ddd, 1H, J2'-3'=4.6
2
Hz, J2'-1'=4.8 Hz, J2'-OH=5.6 Hz, H2'), 5.10 (d, 1H, J3'-OH=5.3 Hz, 3'OH), 5.30 (tq, 1H, JCH-CH2 = 6.6 Hz,
JCH-CH3 = 1.3 Hz, CH=), 5.38 (d, 1H, J2'-OH=5.6 Hz, 2'OH), 5.84 (d, 1H, J1'-2'=4.8 Hz, H1'), 7.77 (br s, 1H,
1
3
NH), 8.21 (s, 1H, H8), 8.29 (s, 1H, H2). C NMR (100 MHz, DMSO-d ): δ = 17.79 (Me), 18.86 (Me),
6
2
5.33 (C5'), 37.67 (NHCH ), 73.05 (C3'), 74.57 (C2'), 79.63 (C4'), 87.84 (C1'), 119.52 (C5), 122.13
2
+
(
CH=), 133.11 (CMe ), 139.53 (C8), 148.43 (C4), 152.53 (C2), 154.30 (C6). HRMS: m/z [M+Na]
2
+
calculated C H N O Na 342.1537, found 342.1534.
1
5
21
5
3
6
4
.5.7. N -(2-phenylethyl)-5′-deoxyadenosine (17)
A mixture of 3 (322 mg, 0.85 mmol), Ph P (334 mg, 1.27 mmol) and 2-phenylethanol (0.153 ml, 1.27
3
mmol) in THF (5 ml) was stirred at ambient temperature until a homogenous solution was formed. DEAD
(0.200 ml, 1.27 mmol) was added in one portion. The reaction was monitored by TLC. After 20 h, a
second addition of reagents (Ph P, 2-phenylethanol and DEAD) in the indicated quantities was made to
3
6
achieve full conversion of initial N -acetyl-2′,3′-di-O-acetyl-5′-deoxyadenosine. After 4-5 h, the mixture
was evaporated and the residue was applied to column chromatography on silica-gel (35 ml). The product
was eluted with methylene chloride:ethanol – 97:3 (100 ml). The fractions, containing the product, were
evaporated in vacuum. The residue was dissolved in 5M propylamine in methanol (8.5 ml) and the
solution was kept for 24 h at 20°С and then concentrated in vacuo to dryness. The residue was evaporated
with ethanol (5×10 ml) and applied to column chromatography on silica-gel (25 ml). The column was
washed with methylene chloride:ethanol – 97:3 (50 ml), the product was eluted with methylene
chloride:ethanol – 9:1. Yield 129 mg (54%) as white powder; R 0.30 (CH Cl -EtOH, 95:5). mp 161-
f
2
2
o
1
1
63 C. H NMR (DMSO-d ): δ = 1.31 (d, 3H, J
= 6.04 Hz, CH ), 2.93 (t, 2H, J
= 7 Hz,
6
CH3-4'
3
CH2-CH2
CH Ph), 3.72 (m, 2H, NHCH ), 3.92-4.02 (m, 2H, H3', H4'), 4.66 (ddd, 1H, J = 4.9 Hz, J2'-3' = 4.5 Hz,
2
2
2'-1'
J2'-OH' = 5 Hz, H2'), 5.10 (d, 1H, J2'-OH = 5.1 Hz, 2'OH), 5.38 (d, 1H, J3'-OH = 5.57 Hz, 3'OH), 5.85 (d, 1H,
1
3
J1'-2' = 4.9 Hz, H1'), 7.15-7.40 (m, 5H, Ph), 7.79 (br s, 1H, NH), 8.24 (s, 1H, H8), 8.30 (s, 1H, H2). C
2
2

ACCEPTED MANUSCRIPT
NMR (DMSO-d ): δ = 18.88 (C5'), 35.01 (NHCH ), 41.25 (CH Ph), 73.07 (C3'), 74.59 (C2'), 79.67 (C4'),
6
2
2
8
7.85 (C1'), 119.54 (C5), 125.98 (Ph), 128.25 (Ph), 128.62 (Ph), 139.62 (C8, Ph), 148.61 (C4), 152.58
+
+
(
C2), 154.50 (C6). HRMS: m/z [M+Na] calculated C H N O Na 378.1537, found 378.1526.
18 21 5 3
5
. Antiviral assay
RD (rhabdomyosarcoma) cells were obtained from the European Collection of Cell Cultures and were
maintained in minimal essential medium (Gibco) supplemented with 10% heat-inactivated fetal bovine
serum (FBS), 1% sodium bicarbonate (Gibco) and 1%
L-glutamine (Gibco). Cells were grown at 37 °C
and 5% CO . EV71 strains that originated from Taiwan were a generous gift from Dr. Shih-Cheng Chang
2
(Chang Gung University, Taoyuan, Taiwan). Strain H08300 461 #812 was acquired from National
Collection of Pathogenic Viruses (NCPV). Strain 11316 was a kind gift from the National Institute for
Public Health and the Environment (RIVM, The Netherlands). EV71 strain BrCr was kindly provided by
F. van Kuppeveld (UMCU, The Netherlands).
The antiviral assay for EV7 was performed as described by Tijsma et al. [28] Briefly, RD cells, grown to
confluence in 96-well microtiter plates were infected with ~100 CCID of EV71 and were treated with a
5
0
dilution series of the different compounds. Cultures were incubated for three days at 37 °C (5% CO2)
after which residual cell viability was quantified using an MTS readout method according to the
manufacturers' instructions (Promega).
CC50 and IC50 values were calculated using a custom-configured software platform from Accelrys.
Acknowledgments
This work was supported by the Russian Foundation for Basic Research (14-04-00835_а and 16-04-
0
1594_a) and Russian Academy of Sciences (Program ‘Molecular and Cell Biology’). Liang Sun is
supported by a CSC: Project of Europe grant from the China Scholarship Council and Aloys Tijsma is
th
supported by the European Union 7 Framework Program EUVIRNA Marie Curie Initial Training
Network (264286). Kim Donckers and Caroline Collared are acknowledged for their excellent assistance
in the acquisition of the antiviral data.
2
3

ACCEPTED MANUSCRIPT
References
[
1] L.Y. Chang, T.Y. Lin, K.H. Hsu, Y.C. Huang, K.L. Lin, C. Hsueh, S.R. Shih, H.C. Ning, M.S.
Hwang, H.S. Wang, C.Y. Lee, Clinical features and risk factors of pulmonary oedema after enterovirus-
1-related hand, foot, and mouth disease, Lancet 354 (1999) 1682–1686.
2] K.T. Wong, B. Munisamy, K.C. Ong, H. Kojima, N. Noriyo, K.B. Chua, B.B. Ong, K. Nagashima,
7
[
The distribution of inflammation and virus in human enterovirus 71 encephalomyelitis suggests possible
viral spread by neural pathways. J. Neuropathol. Exp. Neurol. 67 (2008) 162–169.
[3] W. Xing, Q. Liao, C. Viboud, J. Zhang, J. Sun, J.T. Wu, Z. Chang, F. Liu, V.J. Fang, Y. Zheng, B.J.
Cowling, J.K. Varma, J.J. Farrar, G.M. Leung, H. Yu, Hand, foot, and mouth disease in China, 2008–12:
an epidemiological study, Lancet Infect. Dis. 14 (2014) 308–318.
[
4] P.N. Huang, S.R. Shih, Update on enterovirus 71 infection, Current Opinion in Virology 5 (2014) 98–
04.
5] P. Chong, S.Y. Hsieh, C.C. Liu, A.H. Chou, J.Y. Chang, S.C. Wu, S.J. Liu, Y.H. Chow, I.J. Su, M.
Klein, Production of EV71 vaccine candidates, Human Vaccines and Immunotherapeutics 8 (2012) 1775–
783.
6] L. Shang, M. Xu, Z. Yin, Antiviral drug discovery for the treatment of enterovirus 71 infections,
Antiviral Research 97 (2013) 183–194.
7] H.R. Pourianfar, L. Grollo, Development of antiviral agents toward enterovirus 71 infection, Journal
of Microbiology, Immunology and Infection 48 (2015) 1–8.
8] C.W. Tan, J.K.F. Lai, I.C. Sam, Y.F. Chan, Recent developments in antiviral agents against
enterovirus 71 infection, J. Biomed. Sci. 21 (2014) 1–11.
9] L.Q. Shang, M.Y. Xu, Z. Yin, Antiviral drug discovery for the treatment of enterovirus 71 infections,
Antiviral Res. 97 (2013) 183–194.
1
[
1
[
[
[
[
[
(
[
10] R. L. Kuo, S.R. Shih, Strategies to develop antivirals against enterovirus 71, Virology Journal 10
2013) 28.
11] H. J. Thibaut, A. M. De Palma, J. Neyts, Combating enterovirus replication: State-of-the-art on
antiviral research, Biochem. Pharmacol. 83 (2012) 185–192.
12] M. Arita, T. Wakita, H. Shimizu, Characterization of pharmacologically active compounds that
inhibit poliovirus and enterovirus 71 infectivity, J. Gen. Virol. 89 (2008) 2518–2530.
13] L. Shang, Y. Wang, J. Qing, B. Shu, L. Cao, Z. Lou, P. Gong, Y. Sun, Z. Yin, An adenosine
nucleoside analogue NITD008 inhibits EV71 proliferation. Antiviral Res. 112 (2014) 47–58.
14] V.I. Tararov, A. Tijsma, S.V. Kolyachkina, V.E. Oslovsky, J. Neyts, M.S. Drenichev, P. Leyssen,
[
[
[
S.N. Mikhailov, Chemical modification of the plant isoprenoid cytokinin N6-isopentenyladenosine yields
a selective inhibitor of human enterovirus 71 replication, Eur. J. Med. Chem. 90 (2015) 406–413.
2
4

ACCEPTED MANUSCRIPT
[15] B.B. Mishra, V.K. Tiwari, Natural products: An evolving role in future drug discovery, Eur.[ J. Med.
Chem. 46 (2011) 4769–4807.
16] G.M. Cragg, D.J. Newman, Natural products: A continuing source of novel drug leads, Biochim
Biophys Acta. 1830 (2013) 3670–3695.
17] R. Mah, J. R. Thomas, C. M. Shafer, Drug discovery considerations in the development of covalent
inhibitors, Bioorg Med Chem Lett. 24 (2014) 33–39.
18] J. Zeidler, D. Baraniak, T. Ostrowski, Bioactive nucleoside analogues possessing selected five-
membered azaheterocyclic bases, Eur. J. Med. Chem. 97 (2015) 409-417.
19] Z. Jahnz-Wechmann, G. Framski, P. Januszczyk, J. Boryski, Bioactive fused heterocycles:
Nucleoside analogs with an additional ring, Eur. J Med Chem. 97 (2015) 388-396.
[
[
[
[
6
[
20] V.I. Tararov, S.V. Kolyachkina, C.S. Alexeev, S.N. Mikhailov, N -Acetyl-2',3',5'-tri-O-
6
acetyladenosine – a convenient, missed out substrate for regioselective N -alkylations, Synthesis (2011)
483–2489.
2
[21] M.H. Fleysher, A. Bloch, M.T. Hakala, C.A. Nichol, Synthesis and biological activity of some new
6
N -substituted purine nucleosides, J. Med. Chem. 12 (1969) 1056–1061.
[22] K. Doležal, I. Popa, E. Hauserová, L. Spíchal, K. Chakrabarty, O. Novák, V. Kryštof, J. Voller, J.
6
Holub, M. Strnad, Preparation, biological activity and endogenous occurrence of N -benzyladenosines,
Bioorg. Med. Chem. 15 (2007) 3737–3747.
[
23] S.N. Mikhailov, N.B. Grishko, Acyclic analogs of nucleosides. Synthesis of chiral 1,5-dihydroxy-4-
methyl-3-oxapent-2-yl derivatives of uracyl, Khimiya Geterotsiklicheskikh Soedinenii, (1988) 91–94.
24] M. Ikehara, S. Uesugi, M. Kaneko in: Nucleic Acid Chemistry (L. B. Townsend and R. S. Tipson,
Eds), part 2, p. 610. Wiley, New York 1970.
25] S. Casati, A. Manzocchi, R. Ottria, P. Ciuffreda, H, C and N NMR assignments for N -
isopentenyladenosine/inosine analogues, Magn. Res. Chem. 48 (2010), 745–748.
[
1
13
15
6
[
1
13
15
[
26] J.Vícha, M. Maloň, P Veselá, O. Humpa, M. Strnad, R. Marek, H-, C-, and N-NMR chemical
shifts for selected glucosides and ribosides of aromatic cytokinins, Magn. Res. Chem. 48 (2010) 318–322.
27] S.V. Kolyachkina, V.I. Tararov, C.S. Alexeev, D.M. Krivosheev, G.A. Romanov, E.V. Stepanova,
[
E.S. Solomko, A.N. Inshakov, S.N. Mikhailov, N6-Substituted adenosines. Cytokinin and antitumor
activities, Collect. Czech. Chem. Commun. 76 (2011) 1361–1378.
[28] P. Franchetti, L. Cappellacci, P. Vita, R. Petrelli, A. Lavecchia, S. Kachler, K. N. Klotz, I. Marabese,
L. Luongo, S. Maione, M. Grifantini, N6-Cycloalkyl- and N6-bicycloalkyl-C5'(C2')-modified adenosine
derivatives as high-affinity and selective agonists at the human A1 adenosine receptor with
antinociceptive effects in mice, J. Med. Chem. 52 (2009) 2393-2406.
2
5

ACCEPTED MANUSCRIPT
[29] J. Hsiao, M.-Y. Shen, K-H. Lin, C.-Y. Chou, N.-H. Tzu, C.-H. Lin, D.-S. Chou, T.-F. Chen, J.-R.
Sheu, Inhibitory activity of kinetin on free radical formation of activated platelets in vitro and on
thrombus formation in vivo, Eur. J. Pharmacol. 465 (2003) 281-287.
[30] A. Dassano, M. Mancuso, P. Giardullo, L. De Cecco, P. Ciuffreda, E. Santaniello, A. Saran, T. A.
Dragani, F. Colombo, N6-Isopntenyladenosine and analogs activate the NRF2-mediated antioxidant
response, Redox Biol., 2 (2014) 580-589.
[
31] L. Sáenz, L.H. Jones, C. Oropeza1, D. Vláčil, M. Strnad. Endogenous isoprenoid and aromatic
cytokinins in different plant parts of Cocos nucifera (L.), Plant Growth Regulation 39 (2003) 205–215.
32] P. Mlejnek, P. Dolezel, S. Prochazka. Intracellular conversion of cytokinin bases into corresponding
mononucleotides is related to cell death induction in tobacco BY-2 cells, Plant Science. 168 (2005) 389–
95.
33] Nauš, P.; Pohl, R.; Vortuba, I.; Džubák, P.; Hujdúch, M.; Ameral, R.; Birkuš, G.; Wang, T.; Ray,
[
3
[
A.S.; Mackman, R.; Cihlar, T.; Hocek, M. 6-(Het)aryl-7-deazapurine ribonucleosides as novel potent
cytostatic agents, J. Med. Chem. 53 (2010), 460–470.
[
34] H.E. Gottlieb, V. Kotlyar, A. Nudelman, NMR chemical shifts of common laboratory solvents as
trace impurities, J. Org. Chem. 62 (1977) 7512–7215.
35] P.A. Belyakov, V.I. Kadentsev, A.O. Chizhov, N.G. Kolotyrkina, A.S. Shashkov, V.P. Ananikov,
[
Mechanistic insight into organic and catalytic reactions by joint studies using mass spectrometry and
NMR spectroscopy, Mendeleev Commun. 20 (2010) 125–131.
[
36] L.W. Mclaughlin, N. Piel, T. Hellman, Preparation of protected ribonucleosides suitable for chemical
oligoribonucleotide synthesis, Synthesis (1985) 322–323.
37] S.N. Mikhailov, V.L. Florentiev,W. Pfleiderer. Convenient synthesis of partially blocked oxidized-
reduced nucleosides. Synthesis. 4 (1985) 399-400.
38] A. Mittelman, J. Evans, G. Chheda, Cytokinins as chemotherapeutic agents, Ann. N.Y. Acad. Sci.
55 (1976) 225-234.
[
[
2
2
6

ACCEPTED MANUSCRIPT
6
Modification of the length and structure of the linker of N -benzyladenosine modulates its selective
antiviral activity against enterovirus 71
a1
a1
b
b
a
Mikhail S. Drenichev , Vladimir E. Oslovsky , Liang Sun , Aloys Tijsma , Nikolay N. Kurochkin ,
a
c
b
b
b
Vitali I. Tararov , Alexander O. Chizhov , Johan Neyts , Christophe Pannecouque , Pieter Leyssen ,
Sergey N. Mikhailov *
a
a
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov Str. 32, Moscow
1
19991, Russian Federation
KU Leuven - University of Leuven, Department of Microbiology and Immunology, Laboratory for
b
Virology and Chemotherapy, Rega Institute for Medical Research, Minderbroedersstraat 10, 3000
Leuven, Belgium
c Zelinsky Institute of Organic Chemistry, Russian Academy of Science, Leninsky pr., 47, Moscow
1
19991, Russian Federation
1
Both authors contributed equally.
HIGHLIGHTS
6
N -Benzyladenosine analogs were prepared and tested for antienteroviral activity.
Their activity is greatly dependent on the size and structure of the linker.
6
N -Phenylpropenyladenosine exhibits the highest selectivity index.
The selectivity index was improved in about 50 times.