Design and synthesis of 3,4-dihydro-1H-[1]-benzothieno[2,3-c]pyran and 3,4-dihydro-1H-pyrano[3,4-b]benzofuran derivatives as non-nucleoside inhibitors of HCV NS5B RNA dependent RNA polymerase

Article abstract of DOI:10.1016/j.bmcl.2005.08.114

A novel class of HCV NS5B RNA dependent RNA polymerase inhibitors containing 3,4-dihydro-1H-[1]-benzothieno[2,3-c]pyran and 3,4-dihydro-1H- pyrano[3,4-b]benzofuran scaffolds were designed and synthesized. Optimization of the alkyl substituent in the pyran ring showed preference for an n-propyl group, while 5,8-disubstitution pattern is preferred for the aromatic region. Analog 19 displayed potent activity with an IC₅₀ of 50 nM against HCV NS5B enzyme and was selective over a panel of polymerases.

Full text of DOI:10.1016/j.bmcl.2005.08.114

Bioorganic & Medicinal Chemistry Letters 16 (2006) 457–460
Design and synthesis of
3,4-dihydro-1H-[1]-benzothieno[2,3-c]pyran and 3,4-dihydro-
1H-pyrano[3,4-b]benzofuran derivatives as non-nucleoside
inhibitors of HCV NS5B RNA dependent RNA polymerase
Ariamala Gopalsamy,a,* Alexis Aplasca,a Gregory Ciszewski,a Kaapjoo Park,a
John W. Ellingboe,a Mark Orlowski,b Boris Feldb and Anita Y. M. Howeb
aChemical and Screening Sciences, Wyeth Research, Pearl River, NY 10965, USA
bInfectious Diseases, Wyeth Research, Pearl River, NY 10965, USA
Received 12 July 2005; revised 25 August 2005; accepted 26 August 2005
Available online 7 November 2005
Abstract—A novel class of HCV NS5B RNA dependent RNA polymerase inhibitors containing 3,4-dihydro-1H-[1]-benzothie-
no[2,3-c]pyran and 3,4-dihydro-1H-pyrano[3,4-b]benzofuran scaffolds were designed and synthesized. Optimization of the alkyl sub-
stituent in the pyran ring showed preference for an n-propyl group, while 5,8-disubstitution pattern is preferred for the aromatic
region. Analog 19 displayed potent activity with an IC50 of 50 nM against HCV NS5B enzyme and was selective over a panel of
polymerases.
Ó 2005 Elsevier Ltd. All rights reserved.
Hepatitis C virus (HCV) infection is one of the causes
for liver cirrhosis and hepatocellular carcinoma leading
to liver failure. Current estimates of approximately 170
million people worldwide as HCV carriers represent a
significant medical problem with economic burden
implications.1 Currently approved therapy involves
pegylated interferon-a as a single agent2 or in combina-
tion with the broad spectrum anti-viral ribavirin.2
Although somewhat effective, patient compliance is
compromised due to severe side effects attributed to each
of the agents. Furthermore, the current therapeutic ap-
proach is not aimed at any particular viral target. Lack
of specificity of current therapies against the known
HCV subtypes underscores the need for direct inhibition
of viral targets in an effort to significantly improve pa-
tient outcomes.
tural and non-structural proteins.3 Inhibitors of
NS5B RNA dependent RNA polymerase have re-
ceived much attention recently, as potential therapeu-
tic agents for treatment of HCV infection.4 Both
allosteric and active site inhibitors of NS5B polymer-
ase have been reported (Fig. 1). We recently reported5
pyrano[3,4-b]indole 5 as a potent (IC50 = 0.33 lM) and
selective inhibitor of NS5B polymerase. While the pre-
liminary structure–activity relationship was explored
retaining the pyrano[3,4-b]indole intact, we were inter-
ested in expanding our optimization efforts to some
other scaffolds like 3,4-dihydro-1H-[1]-benzothie-
no[2,3-c]pyran 6 and 3,4-dihydro-1H-pyrano[3,4-b]ben-
zofuran 7 to see if these two different heterocyclic
middle rings, which are devoid of the hydrogen bond
donor capability, are tolerated (Fig. 2).
HCV is a positive strand RNA virus and the genome
consists of 9600 base pairs that encode several struc-
In this communication, we report the synthesis of these
scaffolds and explore the structure–activity relationship
requirements for these new scaffolds.
As shown in Scheme 1, the synthesis of the representative
compound 19 for the 3,4-dihydro-1H-[1]-benzothie-
no[2,3-c]pyran scaffold, started from 1-bromo-2-fluoro-
4-methyl-benzene 8, which upon nitration followed by
Keywords: HCV polymerase inhibitors; 3,4-Dihydro-1H-[1]-benzo
thieno[2,3-c]pyran; 3,4-Dihydro-1H-pyrano[3,4-b]benzofuran.
*
Corresponding author. Fax: +1 845 602 3045; e-mail: gopalsa@
wyeth.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.08.114

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A. Gopalsamy et al. / Bioorg. Med. Chem. Lett. 16 (2006) 457–460
Figure 1. Small molecule inhibitors of HCV NS5B polymerase.
Figure 2. Scaffold substitution for pyrano[3,4-b]indole.
reduction afforded aniline 10. The amino group was con-
verted to the thiol 11 by diazotization followed by reac-
tion with potassium ethyl xanthate. Alkylation of the
thiol followed by deprotection gave the acetic acid deriv-
ative 12, which was converted to the acid chloride 13 and
subjected to Friedel–Crafts cyclization to give the 3-ben-
zothiophenone 14. Wittig reaction of the ketone and sub-
sequent reduction of the ester afforded the alcohol 16.
Treatment of the alcohol 16 with b-ketoester in the pres-
ence of Lewis acid gave the required tricyclic scaffold 17.
The aromatic bromo substituent was converted to cyano
under microwave condition and the ester was hydrolyzed
to give the racemic acid 18 of interest. Further separation
of the required enantiomer was accomplished by chiral
HPLC methods.6
Scheme 1. Reagents and conditions: (a) HNO3, H2SO4, 0 °C, 92%; (b)
SnCl2, EtOAc, 63%; (c) 1—NaNO2, H2O, HCl; 2—potassium ethyl
xanthate; 3–KOH, EtOH, 92%; (d) BrCH2CO2t-Bu, pyridine, 0 °C; (e)
TFA, CH2Cl2, 45% over two steps; (f) SOCl2, 90 °C; (g) AlCl3,
chlorobenzene, 52% over two steps; (h) Ph3P@CHCO2Et, toluene,
120 °C, 39%; (i) LiALH4, THF, 0 °C, 83%; (j) CH3(CH2)2COCH2-
CO2Et, BF3-Et2O, CH2Cl2, 75%; (k) 1—CuCN, NMP, microwave,
220 °C, 15 min; 2—1 N NaOH, EtOH, THF, 61% over two steps; (l)
chiral prep HPLC.6
The synthesis of 3,4-dihydro-1H-pyrano[3,4-b]benzofu-
ran analogs 7 was carried out in an analogous manner
following Scheme 1 where the thiol 11 was replaced with
the appropriately substituted phenol 20 (Scheme 2).
Although, the formation of benzofuran 21 by the cycli-
zation of intermediate phenoxy acetyl chloride proceed-
ed in sub-optimal yield (15–20%), subsequent steps
followed the same trend observed for the 3,4-dihydro-
1H-[1]-benzothieno[2,3-c]pyran scaffold to give the final
tricyclic target 22.
Scheme 2. Reagents and conditions: (m) BrCH2CO2t-Bu, K2CO3,
DMF, rt, 12 h.
methyl to ethyl to n-propyl 23–25, the polymerase inhi-
bition was dramatically improved. However, going to
longer n-butyl group 26 was not favorable indicating
that the n-propyl group is the ideal group for this region
of the molecule.
Moving to the aromatic substitutions, a simple unsubsti-
tuted aromatic system 27, even with the optimal substi-
tution of n-propyl in the C-1 position, was completely
devoid of any activity indicating the need for aromatic
substitutions for the polymerase activity. Addition of
groups like 6-fluoro and 8-methyl (e.g., 28) did not
Our initial efforts were focused on extrapolating the
SAR that we had observed in the case of pyrano[3,4-
b]indole to 3,4-dihydro-1H-[1]-benzothieno[2,3-c]pyran
scaffold. To this end, we started our optimization with
the pyran ring of the molecule. As seen from Table 1,
with changing the alkyl group on the C-1 carbon from

A. Gopalsamy et al. / Bioorg. Med. Chem. Lett. 16 (2006) 457–460
459
Table 1. HCV NS5B inhibitory activity of 3,4-dihydro-1H-[1]-benzo-
thieno[2,3-c]pyran derivatives
than smaller ethyl 36 or methyl 35 analogs and the activ-
ity decreased with longer n-butyl 37 analog. The aromat-
ic substituents were deemed necessary irrespective of the
scaffold employed since compound 34 was inactive.
The most potent inhibitor 19 from this SAR study was
selected for further characterization. Compound 19
showed no inhibitory activity against a panel of human
polymerases9b including mitochondrial DNA polymer-
ase gamma, and other unrelated viral polymerases up
to 50 lM, demonstrating its specificity for the HCV
polymerase. It was found to be selective against helicase
(IC50 = >75 lM) as well as HIV reverse transcriptase
(IC50 = >100 lM). Compound 19 was not cytotoxic in
rapidly dividing and stationary Vero and Huh7 cells as
.
Compound R1
R2
R3 R4
Stereo8 IC50 (lM)9a
23
24
25
26
27
28
29
30
31
32
18
19
33
Me
Et
Cl
Cl
Cl
H
H
H
H
H
F
Cl
Cl
—
—
—
—
—
—
—
—
R
17.6
1.9
n-Pr
Cl
Cl
0.14
0.75
>20
>20
0.19
n-Bu Cl
n-Pr
n-Pr
n-Pr
n-Pr
n-Pr
n-Pr
n-Pr
n-Pr
n-Pr
H
H
Me
Me
Me
Me
Me
Me
Me
Me
H
Br
H
H
H
H
F
measured by
a standard MTS metabolic assay
CN
CN
CN
CN
CN
CN
0.32
0.17
2.85
0.13
0.05
5.36
(IC50 = >75 lM). Moreover, a single administration to
Huh7 cells containing the HCV sub-genomic replicon
for 3 days resulted in a dose-dependent reduction of
the steady-state levels of viral RNA and protein
(EC50 = 3.2 lM for HCV RNA).
S
—
R
F
F
S
In conclusion, we have explored 3,4-dihydro-1H-[1]-
benzothieno[2,3-c]pyran
and
3,4-dihydro-1H-pyr-
satisfy the requirements. However, a 5,8-disubstitution
pattern was found to be regaining substantial amount
of potency as in the case of bromo and cyano derivatives
29 and 30, respectively. A trisubstituted pattern was well
tolerated and showed a slight enhancement in potency.
As observed in the case of pyrano[3,4-b]indole analogs,
one of the enantiomers was more potent than the other
enantiomer as exemplified by the 2 pairs separated 31,
32 and 19, 33.7 It is probably the R enantiomer that
shows activity analogous to the pyrano[3,4-b]indole
series.8
ano[3,4-b]benzofuran derivatives as a novel class of
HCV NS5B RNA dependent RNA polymerase inhibi-
tors. The structure–activity requirement for this class
of inhibitors seems to track very well with the pyr-
ano[3,4-b]indole series. While 3,4-dihydro-1H-[1]-benzo-
thieno[2,3-c]pyran scaffold displayed the same level of
potency as that of pyrano[3,4-b]indoles, 3,4-dihydro-
1H-pyrano[3,4-b]benzofurans were found to be much
less potent. This can probably be attributed to the elec-
tronics of the central rings rather than steric, since the
pyrrole and thiophene rings have a comparable degree
of aromaticity.10 Hence, 3,4-dihydro-1H-[1]-benzothie-
no[2,3-c]pyran scaffold presents itself as a novel chemo-
type specific for HCV polymerase with replicon activity
for further exploration.
Now focusing on the second scaffold of interest, 3,4-
dihydro-1H-pyrano[3,4-b]benzofuran, we synthesized
the proof-of-concept analogs with 5,8-dichloro substitu-
ents in the aromatic region since that pattern showed
good enzyme potency with the 3,4-dihydro-1H-[1]-ben-
zothieno[2,3-c]pyran scaffold. It was interesting to see
that compound 22 (Table 2) was much less potent, when
compared to compound 25. However, the structure–ac-
tivity relationship observed for the C-1 alkyl substitu-
ents showed the same trend with n-propyl being better
Acknowledgments
The authors thank Discovery Analytical Chemistry
group at Wyeth Research, Pearl River, NY, for spectral
data. We thank Drs. Tarek Mansour, John OÕConnell,
David Nunn, and our collaborators at Viropharma,
Inc., for their support of this work and helpful
discussions.
Table 2. HCV NS5B inhibitory activity of 3,4-dihydro-1H-pyrano[3,4-
b]benzofuran derivatives
Supplementary data
Supplementary data associated with this article can be
found in the online version at doi:10.1016/
j.bmcl.2005.08.114.
.
Compound
R1
R2
R4
IC50 (lM)9a
34
35
36
22
37
n-Pr
Me
H
H
>20
>20
16.3
1.49
4.46
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Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Et
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