An efficient synthesis of futalosine

Article abstract of DOI:10.1016/j.tetlet.2010.10.010

Futalosine is a naturally occurring nucleoside comprised of an inosine core with a 3-carboxyphenyl methylene ketone functionality replacing the C-5′ hydroxyl. Recent studies have shown that it is a key intermediate in an alternative biosynthetic pathway that generates menaquinone in a variety of bacterial species. Here we report the first synthesis of futalosine in seven steps from inosine in an overall 17% yield. This work will enable further studies on menaquinone biosynthesis in pathogenic bacteria.

Full text of DOI:10.1016/j.tetlet.2010.10.010

Tetrahedron Letters 51 (2010) 6463–6465
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Tetrahedron Letters
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An efficient synthesis of futalosine
⇑
Xu Li, Martin E. Tanner
University of British Columbia, Department of Chemistry, Vancouver, British Columbia, Canada V6T 1Z1
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 24 July 2010
Accepted 3 October 2010
Futalosine is a naturally occurring nucleoside comprised of an inosine core with a 3-carboxyphenyl meth-
ylene ketone functionality replacing the C-50 hydroxyl. Recent studies have shown that it is a key inter-
mediate in an alternative biosynthetic pathway that generates menaquinone in a variety of bacterial
species. Here we report the first synthesis of futalosine in seven steps from inosine in an overall 17% yield.
This work will enable further studies on menaquinone biosynthesis in pathogenic bacteria.
Ó 2010 Elsevier Ltd. All rights reserved.
Futalosine is a nucleoside that was first isolated in 1999 from
the fermentation broth of Streptomyces sp. MK359-NF1 (Fig. 1).1
It is comprised of an inosine core with a 3-carboxyphenyl methy-
lene ketone functionality replacing the C-50 hydroxyl. More re-
cently, futalosine has been identified as a key intermediate in an
alternative biosynthetic pathway for menaquinone (vitamin
K2).2,3 Studies on Streptomyces coelicolor showed that the men
genes normally responsible for menaquinone biosynthesis in spe-
cies such as Escherichia coli were lacking, and instead this organism
produces the vitamin using a completely novel pathway. Using
gene knockouts it was possible to isolate and identify several of
the intermediates in the pathway, including futalosine. It was dem-
onstrated that futalosine serves as a substrate for futalosine nucle-
osidase (futalosine hydrolase) in both S. coelicolor and Thermus
thermophilus and is converted into dehypoxanthine futalosine
(Fig. 1).4,3 This compound is ultimately converted into menaquin-
one. Bioinformatic analyses indicated that the pathogenic organ-
isms Helicobacter pylori and Campylobacter jejuni also lacked the
men genes, but possessed homologs to the genes of the futalosine
pathway. Since these organisms are the causative agents of duode-
nal ulcers and food poisoning, and since menaquinone biosynthesis
is required for their survival, the futalosine pathway enzymes rep-
resent attractive targets for antibiotic design.5–7 Furthermore, the
absence of the pathway in both humans and beneficial intestinal
bacteria, promises drug treatments with a lack of undesirable side
effects. Further efforts to identify the enzymes of the futalosine
pathway for menaquinone biosynthesis and to develop inhibitors
against them are hampered by the low availability of the sub-
strates. In past studies, modest amounts of futalosine were isolated
from fermentations of mutant strains of S. coelicolor supplemented
with menaquinone.4,3 In order to produce larger amounts of fualo-
sine and to open the doors for analog generation, an efficient syn-
thesis of futalosine was desired. Here we report the first synthesis
of futalosine in seven steps from inosine in an overall 17% yield.
This work will enable further studies on menaquinone biosynthesis
in pathogenic bacteria.
Our first attempt at the synthesis of futalosine involved protec-
tion of the hypoxanthine N1 with a benzyl group and the ribose 2°
hydroxyls as a dimethylacetonide (Scheme 1).
Accordingly, treatment of inosine with benzyl bromide and
NaHCO3 selectively benzylated the N1 position as described previ-
ously.8 Treatment with 2,2-dimethoxypropane and p-toluenesul-
fonic acid then gave compound 1. In order to introduce the
aromatic side chain, a one-pot oxidation/Wittig procedure previ-
ously described by Ikejiri et al. was employed.9 This procedure
avoids the isolation of the C50 aldehyde that can lead to decompo-
sition or epimerization. Phosphorane 2 was first prepared by
O
O
N
NH
N
NH
O
N
N
N
H
N
O
hypoxanthine
O
futalosine
hydrolase
+
OH
O
CO2H
OH OH
futalosine
CO2H
OH OH
dehypoxanthine
futalosine
O
CH3
R
O
menaquinone
R = C40H65
⇑
Corresponding author. Tel.: +1 604 822 9453; fax: +1 604 822 2847.
E-mail address: mtanner@chem.ubc.ca (M.E. Tanner).
Figure 1. The reaction catalyzed by futalosine hydrolase.
0040-4039/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2010.10.010

6464
X. Li, M. E. Tanner / Tetrahedron Letters 51 (2010) 6463–6465
O
N
O
N
N
N
N
N
NH
NH
HO
HO
O
O
1) acetone,
HC(OMe)3,
p-TsOH
1) NaHCO3,
PMB-Cl
2) acetone,
Me2C(OMe)2,
p-TsOH
1) NaHCO3,
BnBr
2) acetone,
Me2C(OMe)2,
p-TsOH
acetone,
HC(OMe)3,
p-TsOH
OH
OH
inosine
OH
inosine
OH
2) POM-Cl, K2CO3
58% (2 steps)
O
O
N
O
N
O
N
N
N
N
NPMB
NPOM
NBn
NH
N
N
N
HO
N
HO
N
N
HO
HO
O
O
O
O
O
O
O
O
O
O
O
PPh3
O
O
O
PPh3
8
11
1
5
2
2
CO2Me
CO2Me
DMP,
phosphorane 2
45%
DMP,
DMP,
phosphorane 2
DMP,
phosphorane 2
phosphorane 2
O
O
N
O
N
O
N
N
N
N
N
NPOM
N
N
NPMB
NH
NBn
O
O
O
O
N
N
N
O
O
O
O
CO2Me
CO2Me
O
O
CO2Me
O
O
CO2Me
O
O
O
O
12
6
9
3
H2,
Pd/C
90%
H2,
H2,
Pd/C
H2,
O
N
O
N
Pd/C
O
N
O
N
Pd(OH)2/C
N
N
N
N
N
NPMB
N
NPOM
NBn
NH
OH
O
O
O
N
N
O
O
O
O
CO2Me
CO2Me
O
O
O
CO2Me
O
O
CO2Me
O
O
O
O
13
O
10
7
4
N
N
NH
N
1) NaOMe, MeOH
2) TFA
3) LiOH
Scheme 1.
O
73% (3 steps)
treatment of m-carbomethoxybenzoyl chloride with methylenetri-
phenylphosphorane.10,11 A mixture of compounds 1 and 2 were
treated with Dess–Martin periodinane (DMP) and gave alkene 3
predominantly as the trans isomer. We then envisioned that reduc-
tion of the alkene and the removal of the benzyl protecting group
could be accomplished in a single step. Treatment of compound 3
with hydrogen and Pd/C cleanly reduced the alkene to give a single
isomer, but the benzyl group remained intact. When the reduction
was carried out with Pearlman0s catalyst, compound 4 was ob-
tained in which both the alkene and the ketone were reduced,
but again the benzyl group remained intact. The difficulty in
removing the N1 benzyl group led us to pursue a synthesis in
which the N1 position of hypoxanthine was left unprotected. Ace-
tonide 5 was prepared by treatment of inosine with acetone, tri-
ethylorthoformate, and p-toluenesulfonic acid as described
previously.12 When acetonide 5 and phosphorane 2 were treated
with DMP, however, a mixture of two isomers of compound 6
was obtained. Hydrogenation of compound 6 also gave an approx-
imately equimolar mixture of isomers of compound 7 that were
inseparable by conventional silica gel chromatography. It would,
therefore, appear that in the absence of a protecting group on
N1, epimerization at C40 occurs during the oxidative coupling
procedure.
CO2H
OH OH
futalosine
Scheme 2.
report describing the advantages of using a p-methoxybenzyl
(PMB) group instead of a benzyl group in protecting inosine,13
prompted us to prepare compound 8 (Scheme 2). The oxidative
coupling of compound 8 with phosphorane 2 produced the trans
isomer of alkene 9. Hydrogenation of the alkene proceeded
smoothly to give a single isomer of compound 10. Unfortunately,
our attempts to remove the N-p-methoxybenzyl group using either
cerric ammonium nitrate or aluminum trichloride/anisole gave low
product yields and/or decomposition. In our final successful syn-
thesis of futalosine, a pivaloyloxymethyl (POM) group was used
to protect the inosine N1.14 The protection of inosine as an aceto-
nide followed by treatment with chloromethyl pivalate and potas-
sium carbonate gave compound 11 in a 58% yield. Oxidative
coupling with phosphorane 2 gave trans alkene 12 in a 45% yield.
Hydrogenation produced compound 13 in a 90% yield. Deprotec-
tion of the POM group proceeded smoothly in sodium methox-
ide/methanol and removal of the acetonide and methyl ester was
achieved using trifluoroacetic acid and lithium hydroxide, respec-
tively. The acidic form of futalosine was generated by treatment
We, therefore, sought to utilize an N1 protecting group that
would be easier to remove at the end of the synthesis. A literature

X. Li, M. E. Tanner / Tetrahedron Letters 51 (2010) 6463–6465
6465
with Amberlite 120H resin (hydrogen form) to give a 73% yield of
References and notes
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identical to those reported for the natural product (see supplemen-
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In summary, a convenient synthesis of futalosine is now avail-
able and can be used to prepare useful amounts of this biosynthetic
intermediate. This work will be useful in enabling studies on men-
aquinone biosynthesis in pathogenic organisms and in the devel-
opment of inhibition assays for the enzyme futalosine hydrolase.
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Acknowledgments
This research was supported by the Canadian Institutes of
Health Research (CIHR).
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Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.tetlet.2010.10.010.