Syntheses of 4′-spirocyclic phosphono-nucleosides as potential inhibitors of hepatitis C virus NS5B polymerase

Article abstract of DOI:10.1016/j.tetlet.2014.06.029

To discover novel nucleosides as potential antiviral agents, 4′-spirocyclic phosphono-nucleosides were designed to mimic the monophosphate of R-1479, a known nucleoside inhibitor of HCV NS5B. Bypassing the first kinase step to nucleoside monophosphate is viewed as advantageous since this phosphorylation is often observed as the rate-limiting transformation to the active NTP for many nucleosides. Efficient synthetic routes were developed with a triphenylphosphine-iodine cyclization reaction as the key step to form the tetrahydrofuran 4′-spirocycle. The desired 4′-spirocyclic phosphono-cytidine analogs 12a, 12b, and 16 were prepared in 11 steps.

Full text of DOI:10.1016/j.tetlet.2014.06.029

Tetrahedron Letters 55 (2014) 4407–4409
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Syntheses of 40-spirocyclic phosphono-nucleosides as potential
inhibitors of hepatitis C virus NS5B polymerase
b
b
b
b
a
Qun Dang a, , Zhibo Zhang , Shuangsheng He , Yaohong Liu , Tongqian Chen , Stephane Bogen ,
⇑
Vinay Girijavallabhan a, David B. Olsen c, Peter T. Meinke a
a Discovery Chemistry, Merck Research Laboratories, 2000 Galloping Hill Road, Kenilworth, NJ 07033, USA
b Pharmaron Beijing Co., Ltd, 6 Tai-He Road, BDA, Beijing 100176, China
c Discovery Biology, Merck Research Laboratories, 770 Sumneytown Pike, West Point, PA 19486, USA
a r t i c l e i n f o
a b s t r a c t
To discover novel nucleosides as potential antiviral agents, 40-spirocyclic phosphono-nucleosides were
designed to mimic the monophosphate of R-1479, a known nucleoside inhibitor of HCV NS5B. Bypassing
the first kinase step to nucleoside monophosphate is viewed as advantageous since this phosphorylation
is often observed as the rate-limiting transformation to the active NTP for many nucleosides. Efficient
synthetic routes were developed with a triphenylphosphine–iodine cyclization reaction as the key step
to form the tetrahydrofuran 40-spirocycle. The desired 40-spirocyclic phosphono-cytidine analogs 12a,
12b, and 16 were prepared in 11 steps.
Article history:
Received 14 May 2014
Revised 4 June 2014
Accepted 5 June 2014
Available online 12 June 2014
Keywords:
Nucleoside
40-Spirocyclic phosphonate
Nucleoside-monophosphate mimic
HCV NS5B inhibitor
Ó 2014 Elsevier Ltd. All rights reserved.
To discover nucleoside inhibitors of HCV NS5B, scientists have
primarily focused on the preparation of nucleoside analogs altered
on the 20-position of the ribose moiety. Representative examples
include 20-modified nucleosides which have successfully advanced
into clinical trials, such as MK-608,1 NM-107,2 PSI-6130,3 and 174
(Fig. 1), although to date only sofosbuvir5 reached the market.
In contrast to the numerous nucleoside analogs with 20-modifi-
cations, R-1479 is a cytidine analog with 40-modification and it is a
potent HCV NS5B inhibitor that advanced to clinical trials (Fig. 2).6
Like all nucleoside inhibitors of HCV NS5B, R-1479 exerts its anti-
HCV activity via its triphosphate (TP) which is the actual inhibitor
of HCV NS5B.7 Thus, R-1479 actually is a prodrug of the active
metabolite R-1479-TP which is formed in vivo via an initial
kinase-mediated phosphorylation step to yield its monophosphate
(R-1479-MP) and then two more kinase transformations to form
the corresponding triphosphate (R-1479-TP). We envisioned that
40-spirocyclic phosphononucleoside 12 could potentially mimic
R-1479-MP as shown in Figure 2.
leading to numerous biologically active molecules with some
reaching the market.8 Attachment of a phosphonate group to the
2-position of the 40-spiro-tetrahydrofuran ring should place
the phosphonate in the same vicinity as the phosphate group of
R-1479-MP. Kinase phosphorylation of the phosphonate 12 could
produce its diphosphate which should mimic R-1479-TP. The suc-
cessful mimicry of the active nucleoside-MPs should bypass the
first kinase step which is often observed as the rate-limiting step
for many nucleosides. It is well established that some organophos-
phonic acids function as substrates for kinases and their
corresponding diphosphates are formed in vivo. For example,
PMEA,9 a phosphonate analog of nucleoside 50-monophosphate,
exerts its anti-HBV activity via its diphosphate and its prodrug
form is employed clinically to treat HBV infections.10
To access the 40-spirocyclicphosphono nucleoside scaffold, a
retrosynthetic plan was devised to prepare the ribose intermediate
8 which should allow rapid exploration of various bases via
glycosidic bond formation reactions, Scheme 1 (Nap: 2-naphthyl-
methyl;11 TBDPS: tert-butyldiphenylsilyl).
The selection of
a phosphonic acid group to mimic the
phosphate of R-1479-MP was based on the observation that
organophosphonates have geometrical and electronic similarities
relative to a phosphate group. Furthermore, organophosphonates
have been demonstrated as excellent mimics of phosphates,
The 40-hydroxymethyl-ribose derivative
1 was prepared
according to the Prakash procedures.12 Treatment of compound 1
with dimethoxytrityl chloride (DMTr-Cl) in pyridine gave the
hydroxy-protected compound 2 in 90% yield. Removal of TBDPS
protecting group using TBAF gave compound 3 in 66% yield, and
subsequent oxidation of the
aldehyde 4 in 95% yield. Wittig reaction of aldehyde 4 gave phos-
a-hydroxymethyl group gave
⇑
Corresponding author. Tel.: +1 908 740 3789.
E-mail address: qun_dang@merck.com (Q. Dang).
http://dx.doi.org/10.1016/j.tetlet.2014.06.029
0040-4039/Ó 2014 Elsevier Ltd. All rights reserved.

4408
Q. Dang et al. / Tetrahedron Letters 55 (2014) 4407–4409
NH2
N
O
O
NH2
N
NH2
N
TBDPSO
DMTrO
TBDPSO
HO
b
d
a
O
O
O
O
O
NapO
NapO
N
1'
N
N
O
N
HO
2
O
1
O
O
HO
O
HO
HO
4'
2'
O
HO OH
MK-608
O
O
HO OH
NM-107
HO
DMTrO
F
c
O
DMTrO
PSI-6130
O
NapO
O
NapO
O
NH2
4
3
NH
N
O
O
O
O
(EtO)2P
HO
N
N
P
O
O
O
O
HN
O
HO
HO
(EtO)2P
O
O
e
O
f
O
O
DMTrO
NapO
O
HO
NapO
F
HO
Sofosbuvir
O
O
O
17
5
6
O
O
Figure 1. Clinically evaluated nucleoside HCV NS5B inhibitors.
(EtO)2P
O
(EtO)2P
O
OAc
OAc
g
O
O
O
NapO
NapO
NH2
N
NH2
N
7
8
O
P
HO
kinases
kinases
R-1479-TP
mimic?
N
Scheme 2. Synthesis of 8. Reagents and conditions: (a) DMTrCl (1.06 equiv),
pyridine, 25 °C, 16 h, 90%; (b) TBAF (2.07 equiv), THF, 25 °C, 48 h, 66%; (c) nBu4NBr
(0.05 equiv), TEMPO (0.05 equiv), KHCO3 (2.1 equiv), NBS (1.57 equiv), CH2Cl2/H2O
(1:1), 0 °C, 1 h, 95%; (d) Ph3P@CH-PO(OEt)2 (1.06 equiv), DMF, 90 °C, 16 h, 65%; (e)
(i) BH3–THF (12 equiv), THF, 25 °C, 12 h; (ii) H2O2 (56 equiv), NaOAc (excess), H2O,
55 °C, 0.5 h, 36%; (f) (i) PPh3 (1.12 equiv), imidazole (2.55 equiv), toluene, reflux,
5 min.; (ii) I2 (1.04 equiv), reflux, 0.5 h, 26%; (g) AcOH/Ac2O/H2SO4 (5.5:2.0:0.2, v/v,
0.08 M solution), 25 °C, 20 h, 52%.
N
HO
O
O
O
O
HO
N3
O
N3
HO OH
R-1479-MP
HO OH
R-1479
NH2
N
O
P
kinases
active DP?
N
HO
HO
O
O
ammonium hydroxide in methanol removed both the N-Ac and 20-
OAc protecting groups to produce 11 in 43% yield, and a final
TMSBr mediated-removal of the phosphonate diethyl ester gave
the desired 40-spirocyclic phosphono nucleosides, which were sep-
arated by HILIC Silica HPLC to give compounds 12a and 12b. The
stereochemistry of the 40-carbon for compounds 12a and 12b
was established via NOESY experiments, and the data are summa-
rized in Figure 3.
O
HO OH
12
Figure 2. Novel 40-spirocyclic phosphono-nucleosides.
phonate ester 5 in 65% yield. Hydroboration of olefin 5 under Negi-
shi conditions13 gave diol 6 in 36% yield (note: the newly formed
chiral center
a
to the phosphorus is racemic; the DMTr protecting
For compound 12a, the 50-methylene (d = 2.42–2.49) exhibited
an NOE interaction with the 30-H (d = 4.28), while the 60-methylene
(d = 3.90–4.10) did not exhibit any NOE interaction with the 30-H
(d = 4.28) and the 6-H (d = 7.77). For compound 12b, the 50-methy-
lene (d = 2.21–2.30) exhibited an NOE interaction with the 6-H
group fell off). Cyclization of the diol 6 to form the tetrahydrofuran
ring was carried out using the Mandal procedure:14 treatment of
diol 6 with triphenylphosphine and iodine in the presence of imid-
azole gave the 40-spiro-tetrahydrofuan 7 in 26% yield (note: the
newly formed chiral center
a to the phosphorus is racemic). Treat-
ment of compound 7 with a mixture of acetic acid, acetic anhy-
dride, and sulfuric acid (5.5:2:0.2, v/v) gave the desired key
ribose derivative 8 (52%), which is ready for Vorbruggen couplings
with various bases (Scheme 2).
With the key ribose 8 in hand, we demonstrated its utility by
coupling with a cytidine base, as illustrated in Scheme 3.
Under Vorbruggen coupling conditions, ribose 8 was reacted
with the silylated N-Ac cytidine in the presence of tin(IV) chloride
to give the nucleoside 9 in 40% yield. Removal of the Nap
protecting group was achieved under DDQ oxidation conditions15
to give compound 10 in 22% yield. Treatment of compound 10 with
NHAc
N
O
O
O
N
(EtO)2P
O
(EtO)2P
O
OAc
a
O
O
NapO
NapO OAc
OAc
8
9
NHAc
NH2
N
N
O
O
O
N
N
(EtO)2P
O
(EtO)2P
O
O
b
c
O
O
OAc
HO
OH
HO
NH2
10
11
O
NH2
NH2
N
N
O
(HO)2P
(EtO)2P
O
N
OAc
OAc
O
N
O
O
(HO)2P
O
O
O
NapO
N
O
N
O
(HO)2P
O
d
O
OH
HO
8
O
O
12
OH
12b
OH
12a
HO
HO
O
TBDPSO
HO
O
Scheme 3. Synthesis of nucleosides 12a and 12b. Reagents and conditions: (a) (i)
N,O-bis(trimethylsilyl)acetamide (7.02 equiv), N-Ac cytidine (2.1 equiv), MeCN,
reflux, 2 h; (ii) compound 8 (1.0 equiv), SnCl4 (3.01 equiv), 25 °C, 15 h, 40%; (b)
DDQ (20 equiv), CH2Cl2/water (20:1, v/v), 25 °C, 48 h, 22%; (c) NH4OH/MeOH (1:1,
v/v), 25 °C, 1.5 h, 43%; (d) TMSBr (15.5 equiv), DMF, 0–25 °C, 4 h, 12a, 8%, 12b, 4.4%.
NapO
O
1
Scheme 1. Retrosynthetic analysis.

Q. Dang et al. / Tetrahedron Letters 55 (2014) 4407–4409
4409
NH2
N
investigation of various bases via glycosidic bond formation reac-
tions. The utility of compound 8 was demonstrated via the synthe-
ses of novel phosphonic acid-containing cytidine nucleoside
analogs 12a, 12b, and 16. Testing of these novel nucleosides
against HCV should provide important SAR information to help
us to expand the understanding of the field.
NH2
N
H
O
H
H
H
6
N
H
H
O H
O
H
N
3' H 1'
4' O
(HO)2P
5'
6'
O
(HO)2P
O
7'O
H
O
H
OH
H
OH
H
OH
OH
12b
12a
Figure 3. Summary of NOE data for compounds 12a and 12b.
Supplementary data
Supplementary data (the experimental procedures and charac-
terization data for all new compounds) associated with this article
can be found, in the online version, at http://dx.doi.org/10.1016/
j.tetlet.2014.06.029.
NHAc
N
F
O
O
(EtO)2P
a
O
OAc
N
(EtO)2P
O
O
O
O
NapO
NapO OAc
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8
13
NHAc
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N
O
c
b
N
(EtO)2P
O
O
O
OAc
HO
14
NH2
N
F
NH2
N
F
O
O
d
N
N
(HO)2P
O
(EtO)2P
O
O
O
O
O
OH
HO
OH
HO
16
15
5. Sofia, M. J.; Bao, D.; Chang, W.; Du, J.; Nagarathnam, D.; Rachakonda, S.; Reddy,
P. G.; Ross, B. S.; Wang, P.; Zhang, H. R.; Bansal, S.; Espiritu, C.; Keilman, M.;
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Leveque, V.; Ma, H.; Maile, G.; Merrett, J. H.; Pichota, A.; Sarma, K.; Smith, M.;
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Scheme 4. Synthesis of nucleosides 16. Reagents and conditions: (a) (i) N-Ac-5-F-
cytidine (2.11 equiv), N,O-bis(trimethylsilyl)acetamide (7.38 equiv), MeCN, reflux,
2 h; (ii) compound 8 (1.0 equiv), SnCl4 (3.16 equiv), 55 °C, 1.5 h, 39%; (b) DDQ
(2.1 equiv), CH2Cl2/water (20:1, v/v), 25 °C, 6 h, 25%; (c) NH4OH/MeOH (1:1, v/v),
25 °C, 15 h, 25%; (d) TMSBr (28.4 equiv), DMF, 0–25 °C, 5 h, 26.
(d = 7.65). On the other hand, the absolute stereochemistry of the
70-carbon was not determined, instead they were assigned arbi-
trarily to differentiate the two diastereo-isomers.
Analogously, 5-fluorocytidine nucleoside 16 was also prepared
from ribose 8 in four steps, as illustrated in Scheme 4.
In summary, to discover active analogs of the known anti-HCV
nucleoside R-1479, a phosphonic acid group was selected as a
phosphate mimic of the key metabolite R-1479-MP. A 40-spirocy-
clic tetrahydrofuran ring was chosen as the linking group which
should provide conformational restriction and partially mimic
the steric effect of the 40-azido group of R-1479. New synthetic
routes have been developed to access a versatile ribose 40-spirocy-
clic phosphonate diester intermediate 8, which should allow rapid
15. Xia, J.; Piskorz, C. F.; Alderfer, J. L.; Locke, R. D.; Matta, K. L. Tetrahedron Lett.
2000, 41, 2773.