Dimeric building blocks with N-cyanoguanidine linkage for oligonucleotide synthesis

Article abstract of DOI:10.1016/S0040-4039(00)60837-7

Synthesis of dimeric nucleoside building blocks with a N-cyanoguanidine linkage was performed by reaction of 3'-amino-3'-deoxythymidine with S,S-dimethyl-N-cyanodithioimidocarbonate followed by reaction of the obtained isothiourea with 5'-amino-2',5'-dideoxynucleosides. These dimers were protected in the 5'-position and converted into their phosphoramidites. Oligonucleotides were synthesized using these building blocks.

Full text of DOI:10.1016/S0040-4039(00)60837-7

Vol. 33. No. 49. pp.
Britain
1992
in
Building Blocks with N-Cyanoguanidine Linkage for
Oligonucleotide Synthesis
C.
P.
Van
and P.
Laboratory of
for Medical Research,
10, B-3000
Belgium
building blocks
a
idinc
by
o f 3 ’ w i t h
b y
t h e o b t a i n e d
in the
and
into
synthesized using these
The synthesis of oligonucleotides with modified nucleosides is of great current interest because of
their potential use as antitumoral and anti-infectious agents (antisense and Natural
constructs suffer from two serious drawbacks their instability against enzymatic degradation and their
poor cellular uptake (and bioavailability) due to their polyanionic character. On the other hand, synthetic
efforts are directed to develop oligonucleotides which are uncharged, enzymatically stable, achiral and
still able to hybridize strongly and selectively with the target
We tried to solve this problem is by synthesizing a.o. oligonucleotides with an
linkage. This functionality is uncharged (at physiological
achiral and enzymatically stable. One idea
which stimulates us to select this substituted guanidine is that the entropy driven free energy change due
to restricted rotation around the intemucleotide linkage during double helix formation would be minimal
starting from a
The synthesis of the
are presented here, were optimalized in order to obtain sufficient material for
rigid
building blocks which we used for oligonucleotide synthesis and which
out
oligonucleotide synthesis on a larger scale. The reactions were first investigated for the synthesis of the
dimer
Reaction of
(1) with
(2)
in ethanol at room temperature gives
(3) which
The cyanoguanidine derivative can be
This
could be isolated directly from the reaction mixture in 80%
obtained from this compound by a substitution reaction with
reaction was carried out in a mixture of triethylamine and dimethylformamide
1.2 equivalent silver nitrate. The yield of this reaction is 80%.
as solvent and using
For the synthesis of the
dimer
no protecting group is needed. Also the

7610
thymine-adenine dimer (6a) can be synthesized without the use of a protecting group. The yield is,
however, better when the group of adenine is protected with a dimethoxytrityl group. Protection
of the base moiety is necessary to obtain good yield of the thymine-cytosine and thymine-guanine dimers.
First the ammo group in the of the cytosine base and the amino group in the of the
base were protected with a
Surprisingly, this protecting group was
level and could not be removed without hydrolysis of the
quite stable at the
function.
A dimethoxytrityl group seemed to be a better choice. Starting from
(4b) and
the thymine-cytosine
thymine-guanine
and
thymine-adenine
dimers were obtained respectively. The dimethoxytrityl protecting groups were
removed with 50% acetic acid for 3 min on a steam bath (6a-c).
This
linkage withstands all protection, deprotection and activation steps necessary
to incorporate them into oligonucleotides. As an example the
the S-position with a dimethoxytrityl group and phosphitylated in the
diisopropylchlorophosphoramidite. This building block (7) was used to synthesize
dimer was protected in
with
(8).
The synthesis of the protected starting materials, the assembly of the dimers to oligonucleotides
and their biological and
properties will be presented in a full paper.
3
1)
2
(3)
A solution of
to a solution of
(5 mmol) of
(15 mmol) of
(1) in 30
ethanol was added
(2) in 10 of
ethanol. The reaction mixture was stirred for 24 h at room temperature. The white precipitate was
collected by filtration giving 1.352 g (4 mmol, 80% yield) of the title compound (3). Elem. anal.
calculated C
H
N 20.64%; found C 45.80 %, H 5.08 %, N 20.58 %.
5
3 vl)
(7)
850 mg (5
of silver nitrate was added to a solution of 1.7 g (5 mmol) of
(3) and 1.3 g (5.4 mmol) of
(4d) in
100
of a mixture of
(1: 1). The reaction mixture was stirred at room temperature for 16 h,
evaporated, coevaporated with xylene and purified by column chromatography
giving 2.13 g (4.01
19.86%.
80% yield) of the title compound (7) as a solid. Elem. anal.
calculated N 20.35%. Found : C :
H
H
N
A. Van Aerschot is a Research Associate of the Belgium National Fund of Scientific Research. We
wish to thank Guy for synthesis of the phosphoramidite. We are indebted to Dominique
Brabants and L. Palmaerts for editorial assistance.

7611
T
T
0
HO
0
HO
2
NC
OH
4
T
T
0
HO
50%
I
B
B
N
0
NC
NC
B ’
T
6
T
T
0
N
NC
P ’
8

7612
Reference&
1. Wickstrom, E. Prospects for Antisense. Nucleic acid therapy of cancer and AIDS.
E.; A. Rev.
3. Williams, D.W. Vol. 24, No. 3,
4. Horwitz, J.-P.; Chua, J.; Noel, M. Chem.
2.
5. Wigerinck, P.,
Aerschot, A, Kerremans, L.,
J., De Clercq, E., Claes, P.
Herdewijn,
P.,
6.
J.P.;
A.J., Urbauski, J.A.,
J. Org. Chem.
7. Nyilas, A.; Glemarec, C.; Chattopadhyaya, J.
8. Gait, M.J. Oligonucleotide synthesis A practical approach; IRL Press
Washington DC,
1984.
(Received in UK 19 August 1992)