A highly stereocontrolled and efficient synthesis of α- and β-pseudouridines

Article abstract of DOI:10.1016/j.tetlet.2003.09.103

A five-step practical and stereocontrolled synthesis of α- and β-pseudouridines from D-ribonolactone is described. The key step involves a highly stereoselective reduction of a hemiketal C-nucleoside intermediate in each case. Multi-gram quantities of β-pseudouridine can now be made available.

Full text of DOI:10.1016/j.tetlet.2003.09.103

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 44 (2003) 8321–8323
A highly stereocontrolled and efficient synthesis of a- and
b-pseudouridines
Stephen Hanessian* and Roger Machaalani
Department of Chemistry, Universite´ de Montre´al, C. P. 6128, Succ. Centre-Ville, Montre´al, P. Q., Canada, H3C 3J7
Received 20 August 2003; accepted 4 September 2003
Abstract—A five-step practical and stereocontrolled synthesis of a- and b-pseudouridines from
D-ribonolactone is described. The
key step involves a highly stereoselective reduction of a hemiketal C-nucleoside intermediate in each case. Multi-gram quantities
of b-pseudouridine can now be made available.
© 2003 Elsevier Ltd. All rights reserved.
The ubiquitous presence of b-pseudouridine 1 (Fig. 1),
a C-5 b-linked uridine in rRNA and tRNA of bacterial
and mammalian origins has instigated numerous studies
concerned with its functional relevance.1 The occur-
rence of pseudouridine in highly conserved regions of
the RNA superstructure reflects on its importance in
molecular recognition phenomena dealing with
polypeptide synthesis for example.2 This and other
observations of biological significance have resulted in
studies aimed at the incorporation of synthetic C-
nucleosides consisting of aromatic and heteroaromatic
aglycones in various RNAs and DNAs.3 The cytotoxic
activity of aromatic C-nucleosides has been known for
some time.4 b-Pseudouridine has been recently reported
as an antimutagenic substance in beer.5
have methods been developed to access C-nucleosides
in preparatively significant amounts.3,4,6 These have
been extended to pyrimidine bases resulting in a practi-
cal 7-step synthesis of b-pseudouridine from readily
available materials such as ribonolactone.6 In spite of
these improvements, a 1:1 mixture of anomeric C-
nucleosides is obtained as a result of a non-selective
reduction of a lactol intermediate with triethylsilane.7
We report herein, a practical, and most importantly, a
highly stereoselective synthesis of b-pseudouridine in
5-steps with an overall yield of 46% starting with
D
-ribonolactone. The method is also applicable to the
synthesis of a-pseudouridine.
Treatment of -ribonolactone
D
3
with
2,2-
As already commented on elsewhere,6 the exploitation
of the potentially important roles that b-pseudouridine
can play in the RNA field has been hampered by its
limited availability and prohibitive cost. Only recently
dimethoxypropane in the presence of pyridinium p-
toluenesulfonate and sodium sulfate gave the protected
bis-acetal 4 in 94% isolated yield (Scheme 1).8 Addition
of 5-lithio-2,4-di-t-butoxypyrimidine 56b to 4 in THF at
−78°C gave the adduct 6 as a 1:8 a/b-mixture in 89%
yield. The latter, in equilibrium with its hydroxyketone
isomer (not shown) was reduced with
L-Selectride (3.4
equiv.) in the presence of ZnCl2 (1.36 equiv.) to give the
acyclic -altro-hexitol 7 as a single isomer in 85% yield.
D
Cycloetherification under Mitsunobu conditions9 with
diisopropyl azodicarboxylate (DIAD) and triphenyl-
phosphine gave 8 in 70% yield. Finally, deprotection
with 70% acetic acid gave b-pseudouridine in 93% yield,
identical with an authentic sample. This constitutes the
shortest synthesis of b-pseudouridine which can now be
prepared in multi-gram quantities.10
Figure 1. Structures of a- and b-pseudouridine.
The anomeric a-pseudouridine 2,6b was synthesized
essentially using the same protocol, except that the
Keywords: C-nucleoside; cycloetherification; selective reductions.
* Corresponding author.
0040-4039/$ - see front matter © 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2003.09.103

8322
S. Hanessian, R. Machaalani / Tetrahedron Letters 44 (2003) 8321–8323
Scheme 2.
as described above gave a-pseudouridine 2 with an
overall yield of 66%. It is of interest that the Mitsunobu
cyclization was faster and more efficient in the case of
9 compared to 7, probably due to a lower steric
barrier.11
In an earlier version of the synthesis where 2,4-
dimethoxypyrimidine was the aglycone, hydrolysis with
NaI/TMSCl (MeCN, 16 h) according to a literature
procedure12 (or with 70% AcOH, 30% HBr in AcOH,
reflux 1 h) led to considerable anomerization. Related
epimerizations have been recently reported for aromatic
C-nucleosides under mild acidic conditions.13 A plausi-
ble mechanism is shown in Scheme 3.
Scheme 1.
reduction of the lactol intermediate 6 was done with
L
-Selectride in the absence of zinc chloride (Scheme 2).
Remarkably, an >20:1 a/b-mixture of the
D
-allo-9 and
D
-altro-7 hexitols respectively was obtained in 88%
yield. Subsequent cycloetherification and deprotection
Scheme 3.
Figure 2. Possible pathways for stereoselective reductions.

S. Hanessian, R. Machaalani / Tetrahedron Letters 44 (2003) 8321–8323
8323
The remarkably stereoselective reduction of the lactol 6
with -Selectride in the presence or absence of zinc
chloride can be rationalized by invoking chelated and
non-chelated intermediates, respectively as illustrated in
Figure 2. Chelation of the divalent zinc with the ketone
carbonyl and the acetal oxygen allows hydride delivery
67, 3724; for reviews, see (b) Mitsunobu, O. Synthesis
1981, 1; (c) Hughes, D. C. Org. React. 1992, 42, 335.
10. Typical procedure 6“8 for b-pseudouridine.
L
7: To 6 (450 mg, 0.93 mmol) in dry DCM (36 mL) at
−78°C, was added ZnCl2 (1.26 mL, 1.26 mmol, 1 M in
Et2O) and the mixture was stirred for 30 min. L-Selectride
from the Si (pro-S) face to give the observed
D-altro
(3.16 mL, 3.16 mmol, 1 M in THF) was then added
dropwise and portionwise for a period of 30 min at −78°.
Mixture was then slowly brought to room temperature
and stirred overnight. The white heterogeneous solution
was quenched with EtOH (0.54 mL), H2O (0.120 mL),
NaOH 6M (0.47 mL) and H2O2 30% (0.47 mL). It was
then stirred for 30 min and diluted with EtOAc (50 mL)
and H2O (50 mL). Usual workup gave a colorless oil
which was purified by flash chromatography (1:1 hex-
anes/EtOAc) to give 7 as a white foamy solid (385 mg,
85%); Rf=0.35 (1:1 hexanes/EtOAc), m.p. 40–42°C, [h]D
−50.64° (c 0.23, MeOH). 8: To 7 (155 mg, 0.32 mmol) in
dry THF (33 mL) was added Ph3P (168 mg, 0.64 mmol).
The colorless mixture was cooled to 0°C and DIAD
(0.126 mL, 0.64 mmol) was added, stirred at 4°C for 48 h,
then brought to room temperature and concentrated. The
yellow oil was purified by flash chromatography (2:1
hexanes/EtOAc) to give 8 as a colorless oil (105 mg,
70%); Rf=0.72 (1:1 hexanes/EtOAc); [h]D +12.0° (c 8.5,
MeOH).
isomer 7. In the absence of chelation, the Re(pro-R)-
face of the carbonyl is better exposed to receive the
bulky hydride. The influence of the protective groups
on the heterocycle (ex. 2-C-benzimidazolyl; 2-C-imida-
zolyl, etc.) on the selectivity of reductions of C-hetero-
cyclic lactols with NaBH4, has been reported.9a
The extension of this new method to the stereoselective
synthesis of other C-nucleosides is currently in
progress.
Acknowledgements
We thank NSERC, Syngenta Crop Protection (UK) for
generous financial support through the Medicinal
Chemistry Chair Program.
11. Typical procedure 6“9“10 for a-pseudouridine.
9: To 6 (500 mg, 1.03 mmol) in dry THF (40 mL) at
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