Synthesis of tritium-labeled 2′,3′-dideoxy-2′,3′- didehydrothymidine

Full text of DOI:10.1002/jlcr.1287

Journal of Labelled Compounds and Radiopharmaceuticals
J Label Compd Radiopharm 2007; 50: 585–587.
Published online in Wiley InterScience
JLCR
(www.interscience.wiley.com). DOI: 10.1002/jlcr.1287
Short Research Article
Synthesis of tritium-labeled 20,30-dideoxy-20,30-
didehydrothymidiney
G. V. SIDOROV1,*, N. F. MYASOEDOV1, M. V. JASKO2 and YU. S. SKOBLOV2
1 Institute of Molecular Genetics, Russian Academy of Sciences, Kurchatov Sq. 2, Moscow 123182, Russia
2 Institute of Molecular Biology, Russian Academy of Sciences, Vavilov St. 32, Moscow 119991, Russia
Received 6 July 2006; Revised 22 January 2007; Accepted 24 January 2007
Keywords: tritium; isotope-exchange; 20,30-dideoxy-20,30-didehydrothymidine
Introduction
Results and discussion
The mechanisms of action, metabolic pathways, and
cellular uptake of 20,30-dideoxy-20,30-didehydrothymi-
dine have been intensively studied in recent years.1
These studies require tritium-labeled compound at
high specific radioactivity ðAmol > 10 Ci=mmolÞ. A new
method of synthesis of tritium-labeled 20,30-dideoxy-
20,30-didehydrothymidine was developed. The first
stage was the preparation of tritium-labeled 50-O-
benzoyl-2,30-anhydrothymidine by catalytic isotope
exchange with gaseous tritium in solution. The synthe-
sized [6-3H]50-O-benzoyl-2,30-anhydrothymidine was
Synthesis was carried out according to Scheme 1:
In the first step we synthesized [6-3H]50-O-benzoyl-
2,30-anhydrothymidine. Previous NMR studies had
shown that tritium replaces position 6 of pyrimidine.2
The second step was an elimination reaction resulting
in the formation of [6-3H]20,30-dideoxy-20,30-didehy-
drothymidine. During this step the O-benzoyl residue
was removed quantitatively. Most likely excess of NaH
causes degradation the DMF with the formation of
dimethylamine, which removes O-benzoyl group. A
‘cold’ synthesis of 20,30-dideoxy-20,30-didehydrothymi-
dine was also carried out and the NMR spectrum
(Figure 1) confirmed the formation of 20,30-dideoxy-
20,30-didehydrothymidine.
transformed
into
[6-3H]20,30-dideoxy-20,30-didehy-
drothymidine in one stage, in an elimination reaction
catalysed by sodium hydride in DMF.
O
O
O
N
CH3
3H2, Pd/CaCO3
CH3
3H
HN
N
CH3
3H
N
N
N
O
NaH
O
O
HO
BzO
BzO
O
O
0.1 N NH3/H2O
O
DMF
(III)
(II)
(I)
Scheme 1
Synthesis of tritium-labeled [6-3H]50-O-benzoyl-2,30-
anhydrothymidine (II)
*Correspondence to: G. V. Sidorov, Institute of Molecular Genetics,
Russian Academy of Science, Kurchatov Sq. 2, Moscow 123182,
Russia. E-mail: sidgv@img.ras.ru
Contract/grant sponsor: Russian Foundation for Basic Research;
contract/grant numbers: 05-04-49500, 04-03-32481
yProceedings of the Ninth International Symposium on the Synthesis
and Applications of Isotopically Labelled Compounds, Edinburgh,
16–20 July 2006.
50-O-Benzoyl-2,30-anhydrothymidine ðIÞ (6.5 mg, 19.8
mmol) was placed in a reaction glass reaction vial, then
500 ml of 0.1 N ammonia in 50% aqueous ethanol and
90 mg of 5% Pd/CaCO3 (Fluka) as catalyst were added.
The vial was then frozen in liquid nitrogen, evacuated,
Copyright # 2007 John Wiley & Sons, Ltd.

586 G. V. SIDOROV ET AL.
Figure 1 NMR spectrum of 20,30-dideoxy-20,30-didehydrothymidine (‘cold’ synthesis) in D2O. AMX III-400 (Bruker) spectrometer
with the 400 MHz operating frequency for 1H.
and filled with gaseous tritium to
a
pressure of
was achieved by HPLC. As a result, 17 mCi of product
was obtained with a specific radioactivity of 18.5 Ci/
mmol, in 20% yield. The radiochemical purity of the
product as estimated by thin layer chromatography
was above 97%. [6-3H]20,30-dideoxy-20,30-didehy-
drothymidine was stored at ꢁ208C as a solution of
1 mCi/ml in 1:1 (v/v) water–ethanol.
400 mmHg. Upon defrosting, the reaction mixture was
stirred at room temperature for 20 h. After the reaction
was completed, tritium was removed from the vial and
the catalyst separated by centrifugation. Labile tritium
was removed by evaporation with 10 ml of 50% ethanol.
Isolation of the product was achieved by HPLC. As a
result, 170 mCi of product was obtained with a specific
radioactivity of 21.6 Ci/mmol, in 38% yield.
HPLC was conducted using a 13-mm ð10 ꢀ 250 mmÞ
Nucleosil C18 (Macherey–Nagel) column, elution rate
1.5 ml/min, detection UV 260 nm. The mobile phase:
50-O-Benzoyl-2,30-anhydrothymidine. 60% Methanol
*
Synthesis of tritium-labeled 20,30-dideoxy-20,30-dide-
hydrothymidine (III)
in water, retention time 13.1 min.
20,30-Dideoxy-20,30-didehydrothymidine. Solvents: A
*
[6-3H]50-O-benzoyl-2,30-anhydrothymidine ðIIÞ (1.3 mg,
85 mCi) was dissolved in 0.1 ml of absolute
N,N-dimethylformamide, then about 1 mg of NaH
(80% in oil) was added and the solution was stirred at
room temperature for 18 h. Then 10 ml of 10% acetic
acid was added. The solution was evaporated in
vacuum, co-evaporated with water ð4 ꢀ 0:2 mlÞ and
then with ethanol ð2 ꢀ 0:4 mlÞ. Isolation of the product
– water; B – methanol. Program: 0–40% B/A in
40 min, retention time 33.9 min.
TLC was carried out on Silufol plates (Czech Republic).
50-O-Benzoyl-2,30-anhydrothymidine in chloroform–
*
ethanol (95:5 v/v) solvent system, Rf ¼ 0:35.
20,30-Dideoxy-20,30-didehydrothymidine chloroform–
*
ethanol (9:1 v/v) solvent system, Rf ¼ 0:28.
Copyright # 2007 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2007; 50: 585–587
DOI: 10.1002.jlcr

SYNTHESIS OF 20,30-DIDEHYDROTHYMIDINE 587
Acknowledgements
REFERENCES
The work was supported by the Russian Foundation for
1. De Clercq E. Nat Rev Microbiol 2004; 2: 704.
2. Sidorov GV, Zverkov YuB, Myasoedov NF, Jasko MV,
Skoblov YuS. J Label Compd Radiopharm 2003; 46: 669.
Basic
Research,
projects
05-04-49500
and
04-03-32481.
Copyright # 2007 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2007; 50: 585–587
DOI: 10.1002.jlcr