Hepatitis C NS5B polymerase inhibitors: 4,4-Dialkyl-1-hydroxy-3-oxo-3,4-dihydronaphthalene-3-yl benzothiadiazine derivatives

Article abstract of DOI:10.1016/j.bmcl.2008.06.043

4,4-Dialkyl-1-hydroxy-3-oxo-3.4-dihydronaphthalene-3-yl benzothiadiazine derivatives were synthesized and evaluated as inhibitors of genotypes 1a and 1b HCV NS5B polymerase. A number of these compounds exhibited potent activity against genotypes 1a and 1b HCV polymerase in both enzymatic and cell culture activities. A representative compound also showed favorable pharmacokinetics in the rat.

Full text of DOI:10.1016/j.bmcl.2008.06.043

Bioorganic & Medicinal Chemistry Letters 18 (2008) 3887–3890
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Hepatitis C NS5B polymerase inhibitors: 4,4-Dialkyl-1-hydroxy-3-oxo-3,
4-dihydronaphthalene-3-yl benzothiadiazine derivatives
a
a
a
a
Douglas K. Hutchinson a, , Teresa Rosenberg , Larry L. Klein , Todd D. Bosse , Daniel P. Larson ,
Wenping He a, Wen W. Jiang a, Warren M. Kati a, William E. Kohlbrenner a, Yaya Liu a,
Sherie V. Masse a, Tim Middleton a, Akhteruzzaman Molla a, Debra A. Montgomery a,
David W. A. Beno b, Kent D. Stewart c, Vincent S. Stoll c, Dale J. Kempf a
*
a Department of Antiviral Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064, USA
b Department of Drug Metabolism and Pharmacokinetics, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064, USA
c Department of Structural Biology, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
4,4-Dialkyl-1-hydroxy-3-oxo-3.4-dihydronaphthalene-3-yl benzothiadiazine derivatives were synthe-
sized and evaluated as inhibitors of genotypes 1a and 1b HCV NS5B polymerase. A number of these com-
pounds exhibited potent activity against genotypes 1a and 1b HCV polymerase in both enzymatic and cell
culture activities. A representative compound also showed favorable pharmacokinetics in the rat.
Ó 2008 Elsevier Ltd. All rights reserved.
Received 31 March 2008
Revised 10 June 2008
Accepted 12 June 2008
Available online 18 June 2008
Keywords:
Hepatitis C
HCV
HCV NS5B polymerase
Antiviral
Hepatitis C is a positive-stranded RNA virus of the Flaviviridae
family which was first identified in 1989.1 Since that time, the hep-
atitis C virus (HCV) has been found to be a major cause of cirrhosis,
hepatocellular carcinoma, as well as liver failure, and is the most
common cause in developed countries of the need for liver trans-
plantation.2 HCV is most commonly transmitted by blood transfu-
sions, hemodialysis, and intravenous drug use, and is quite
insidious in that clinical manifestations can be mild or non-extant
for years or even decades after initial infection. It is estimated by
the Centers for Disease Control (CDC) that 2.7 million Americans
and a total of 170 million individuals worldwide are chronically in-
fected with HCV, this number being approximately 3% of the world
population.3 In addition, the CDC estimates that 8–10 thousand
deaths each year in the United States alone are related to HCV,
and this number is expected to triple over the next two decades.
It can be seen that clinical manifestations of HCV infection rep-
resent a major impending healthcare problem. Unfortunately, the
genotypes 1 and 2 are the most common in the United States, Eur-
ope, and Japan.6 Sustained virologic response (SVR) in patients in-
fected with HCV genotypes 2 and 3 approaches 80% after a 24-
week treatment regimen of PEG-IFN-a/ribavarin. In contrast, SVR
in patients infected with genotype 1 is less than 50% even after ex-
tended treatment for 48 weeks.4,5 Consequently there is a great
medical need for new HCV chemotherapeutic agents that would
give SVR against genotypes other than 2 or 3.
The cloning of HCV in 19891 eventually led to identification of
several targets for small molecule intervention in the virus life cy-
cle.7,8 Of these, inhibitors of the NS3/NS4A protease/helicase and
the NS5B RNA-dependent RNA polymerase (RdRp) have been the
most actively investigated.8 Drug discovery efforts aimed at inhib-
iting the HCV NS5B RdRp have disclosed both nucleoside and non-
nucleoside inhibitors.8 One series of non-nucleoside inhibitors that
show promise in both enzymatic and cell-based replication assays
using the subgenomic HCV replicon are the benzothiadiazines, of
which structure 1 is representative.9 This series of compounds,
originally prepared as diuretics,10 was found to be potent inhibi-
tors of HCV NS5B polymerase in a manner noncompetitive with re-
spect to GDP.10 Mechanistically these compounds were found to
inhibit initiation of viral RNA synthesis by preventing the binding
of the RNA primer to the enzyme11 (Fig. 1).
current treatment regimens utilizing pegylated interferon-a-2a
(PEG-IFN-a
, PEG-IntronÒ) in combination with ribavarin (Rebe-
trolÒ and CopegusÒ) are extremely expensive and are accompanied
by serious systemic side effects.4,5 This is complicated by the exis-
tence of six major genotypes of HCV (genotypes 1–6), of which
As part of ongoing drug discovery efforts at Abbott, hydrazine
derivatives, such as 2, were found to be potent inhibitors of HCV
* Corresponding author. Tel.: +1 847 938 8441; fax: +1 847 938 2756.
E-mail address: douglas.hutchinson@abbott.com (D.K. Hutchinson).
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.06.043

3888
D. K. Hutchinson et al. / Bioorg. Med. Chem. Lett. 18 (2008) 3887–3890
diethyl malonate under the conditions of Rathke17 afforded ketodi-
ester 9 in good overall yield. Cyclization of 9 was readily accom-
plished by stirring in methanesulfonic acid overnight, affording
diketo ester 10 in good yield. After hydrolysis/decarboxylation,
resultant diketone 11 was treated with salt 13 to afford dithioke-
tene acetal 12 in moderate to good yields.18
Precursors to the benzothiadiazine ring system were synthe-
sized by methods summarized below in Scheme 2. As shown, ami-
nation of benzenesulfonamide 14 afforded 15 in 90% yield.19
Reduction of the nitro group and treatment with methanesulfonyl
chloride gave the key methanesulfonamide 16. For other substitit-
uents on the benzothiadiazine ring system, synthetic intermedi-
ates were prepared by modifications of methods developed by
Topliss.20 As shown, benzylation of nitrophenol 17 followed by
nucleophilic aromatic displacement with benzyl mercaptan, affor-
ded thioether 20 in excellent overall yield. Oxidation of 20 with
chlorine gas and then quenching the intermediate sulfonyl chloride
with aqueous ammonia, afforded the nitro benzenesulfonamide 21
in 89% overall yield. Reduction of the nitro group with iron in the
presence of ammonium chloride afforded aniline 22 in good
yield.
O
O
O
O
S
S
OH
N
OH
N
N
N
H
N
H
N
NH
O
O
2
1
O
O
S
O
O
OH
N
S
OH
N
N
H
N
H
O
O
R
R
4
3
Figure 1. Genesis of dialkyl benzothiadiazines.
polymerase both in enzymatic and cell-based replication assays.12
While some analogs of 2 exhibited good pharmacological proper-
ties, others showed extremely poor solubility and were difficult
to formulate for in vivo studies. Postulating that replacement of
the nitrogen atom of the dihydroquinolinedione ring system of 1
with a carbon atom, as in structure 3, would interfere with the
Final assembly and functional group interconversions of ana-
logs for biological evaluation are summarized in Scheme 3. As
shown, treatment of ketenedithioacetal 12 with amino sulfon-
amide 22 in refluxing dioxane afforded benzothiadiazine 23 in
good yield. Compound 23 was then transformed into the oxyaceta-
mide analog 25 in excellent overall yield using routine chemical
transformations. In a similar manner, treatment of 12 with 16
afforded methanesulfonamide 26 in good yield.
A number of structurally diverse compounds synthesized in an
analogous manner were evaluated in biochemical assays against
HCV NS5B polymerases from genotypes 1a and 1b. In addition,
the activities of these compounds in cell culture against the HCV
subgenomic replicons from genotypes 1a and 1b were also deter-
mined. These data are summarized in Table 1 below (see Supple-
mentary material for biological methods).
compound’s
p-stacking ability, it would follow that such com-
pound would be less crystalline, and therefore more soluble. A sim-
ilar approach was used by Rosen to improve the solubility of a
series of fluoroquinolone antibacterial agents.13 Preparation of 3
would, however, necessitate an asymmetric synthesis or a resolu-
tion, the difficulty of which would not be justified for a speculative
series of analogs. Consequently, the preparation of a symmetrically
disubstituted analog series, illustrated by structure 4, was under-
taken for proof of concept.
As shown, a wide range of activities were observed for the gem-
dialkyl benzothiadiazines. Based on published crystal structure
data of structurally related inhibitors in the benzothiadiazine ser-
ies, it can be postulated that the gem-dimethyl analogs (27 and
28) would show, at best, modest levels of activities, since they lack
a longer hydrocarbon chain to project into the hydrophobic pocket
below the plane of the ring systems.21 Consistent with this theory,
increasing the chain length improved the potency, with gem-dipro-
pyl methanesulfonamide 31 showing a nearly 10-fold increase in
activity over its gem-dimethyl counterpart 28. In most cases exam-
The synthesis of the 4,4-dialkyl-1-hydroxy-3-oxo-3.4-dihydro-
naphthalene ring system was easily accomplished by a modifica-
tion of the method of Buckle.14 The synthesis of the gem-dibutyl
series was typical of that used for all analog series investigated,
and is summarized in Scheme 1. As shown, alkylation of methyl
phenylacetate (5) with crotyl bromide afforded the di-(2-butenyl)
compound 6 in high yield. After hydrogenation of the double bonds
and hydrolysis of the ester,12,15 the acid 8 was treated with an ex-
cess of oxalyl chloride in the presence one equivalent of DMF13,16
to cleanly afford the acid chloride. Subsequent treatment with
a
b
c
O
O
O
O
O
OH
O
O
5
8
6
7
-
BF4
S+
d, e
S
S
O
OH
O
O
OH
O
S
O
O
13
g
O
f
S
O
h
O
O
O
10
9
11
12
Scheme 1. Reagents and conditions: (a) crotyl bromide, NaH, NaI, THF,
(3 equiv), DMF (1 equiv), hexanes; (e) diethyl malonate, MgCl2, TEA, CH3CN, 95% overall; (f) methanesulfonic acid, RT, 60%; (g) 1:1 1 N HCl–THF,
dioxane, , 70%.
D
, 95%; (b) H2 (45 psi), 10% Pd/C, MeOH, 95%; (c) KOSiMe3 (5 equiv), THF,
D
, 90%; (d) oxalyl chloride
, 70%; (h) 13, pyridine,
D
D

D. K. Hutchinson et al. / Bioorg. Med. Chem. Lett. 18 (2008) 3887–3890
3889
O O
S
O O
S
O O
S
H
N
NH2
NO2
a, b
H2N
c
H2N
S
H2N
O O
H2N
H2N
Cl
14
15
16
F
O
Ph
Ph
S
O
Ph
F
OH
d
e
O2N
O2N
O2N
17
18
20
f
O O
O O
S
S
O
Ph
O
Ph
g
H2N
O2N
H2N
H2N
21
22
Scheme 2. Reagents and conditions: (a) (NH4)2CO3, CuSO4, NH4OH, 120 °C, sealed tube, 90%; (b) Na2S2O4, NaOH, 70%; (c) MsCl, pyridine, CH2Cl2, 60%; (d) benzyl bromide,
Cs2CO3, TBAI, DMF, 100%; (e) benzyl mercaptan, Na2CO3, EtOH,
D
, 90%; (f) i—Cl2, HOAc; 0 °C; ii—NH4OH, CH2Cl2, 89% overall; (g) Fe, NH4Cl, MeOH–H2O, D, 72%.
O
O
S
O
Ph
O
S
O
O
H2N
H2N
O
OH
S
O
Ph
OH
N
OH
N
O
S
22
b
N
H
N
H
S
a
O
O
O
24
23
12
O
O
H
N
S
O
O
O
O
H2N
H2N
S
O
H
N
c
S
S
O
O
O
OH
N
S
OH
N
NH2
O
O
16
N
H
N
H
a
O
O
26
25
Scheme 3. Reagents and conditions: (a) dioxane, 100 °C, 75%; (b) H2, 10% Pd/C, EtOAc, 90%, (c) ICH2CONH2, TBAI, Cs2CO3, DMF, 90%.
ined, the methanesulfonamide substituent on the D ring gave 2- to
4-fold better activities than those obtained using the oxyacetamide
substituent. The gem-dibutyl series showed the best overall in vitro
activity in this series, with gem-dibutyl oxyacetamide derivative 25
and methanesulfonamide derivative 26 showing submicromolar
IC50 values of 10 and 73 nM against HCV strains 1a and 1b, respec-
tively. In addition, gem-dibutyl methanesulfonamide 26 also
showed submicromolar levels of activity in the replicon assays,
against both the 1a and 1b strains. While the iso-amyl side chain
had afforded potent analogs in previous series, in this case the
gem-iso-amyl compounds 34 and 35 showed less activity than their
n-butyl counterparts, possibly because of the increased energy cost
of displacing more water molecules from the space above the plane
of the rings at the binding site. Consequently, while the iso-amyl
group may be ideal for binding in the hydrophobic pocket in other
analog series,21 the n-butyl group may represent a compromise be-
tween binding affinity and the energy cost of displacing waters
from the binding site.
The pharmacokinetic behavior of gem-dibutyl methanesulfon-
amide 26 in the rat was examined using both intravenous and oral
doses of 5 mg/kg. As shown in Table 2, the compound shows low
clearance with a moderate volume of distribution. With oral dos-
ing, the compound shows a half-life of 4.3 h, with a maximum plas-
ma concentration of 0.58 lg/mL occurring at 3 h. With exposure
levels obtained from oral dosing being similar to those obtained
from intravenous dosing, these data correspond to a bioavailability
of 94%.
The pharmacokinetic behavior of 26 was even more interesting
when liver concentrations were determined. As shown in Table 3,
much higher levels of 26 were found in the liver than in plasma,
up to 12 h after dosing. Maximum liver concentrations of 21
lg/g
were achieved 6 h after dosing, whereas a 0.41- g/mL concentra-
l
tion was achieved in the plasma, this corresponding to a 48:1 liver
to plasma ratio.
In conclusion, it can be stated that while most of the com-
pounds in this study had only modest activity, the gem-dibutyl

3890
D. K. Hutchinson et al. / Bioorg. Med. Chem. Lett. 18 (2008) 3887–3890
Table 1
Biochemical and cell culture activity of gem-dialkyl benzothiadiazines
O O
S
C
R2
OH
N
D
N
A
B
H
O
R1 R1
Compound
R1
R2
Polymerase 1a IC50
(l
M)a
Polymerase 1b IC50
(l
M)a
Replicon 1a EC50
(l
M)a,b
Replicon 1b EC50 (l
M)a,b
27
28
29
30
31
32
25
26
33
34
35
1
CH3
CH3
H
2.2
—
6.26
—
—
—
—
—
—
—
NHSO2CH3
H
OCH2CONH2
NHSO2CH3
H
OCH2CONH2
NHSO2CH3
H
OCH2CONH2
NHSO2CH3
0.93
2.20
0.24
0.099
0.99
0.26
0.010
0.82
0.32
0.068
0.54
0.28
n-Propyl
n-Propyl
n-Propyl
n-Butyl
n-Butyl
n-Butyl
iso-Amyl
iso-Amyl
iso-Amyl
—
>10.0
0.60
—
9.64
0.28
—
10.10
0.42
—
0.81
0.022
—
0.29
0.048
—
>3.0
—
0.25
0.10
0.13
2.27
0.33
0.073
—
—
0.39
0.42
0.48
2
—
a
Both IC50 and EC50 values are means of at least two independent determinations, standard deviation 10 %. Detailed protocols can be found in Supplementary material.
Assay run with 5 % fetal calf serum.
b
2. Poordad, F. F.; Tran, T.; Martin, P. Expert Opin. Emerging Drugs 2003, 8, 9.
Table 2
3. Alter, M. J.; Kruszon-Moran, D.; Nainan, O. V.; McQuillan, G. M.; Gao, F.; Moyer,
L. A.; Kaslow, R. A.; Margolis, H. S. N. Engl. J. Med. 1999, 341, 556.
4. Manns, M. P.; McHutchison, J. G.; Gordon, S. C.; Rustgi, V. K.; Shiffman, M.;
Reindollar, R.; Goodman, Z. D.; Koury, K.; Ling, M.-H.; Albrecht, J. K. Lancet
2002, 347, 975.
5. Fried, M. W.; Shiffman, M. L.; Reddy, K. R.; Smith, C.; Marinos, G.; Goncales, F. L.,
Jr.; Haussinger, D.; Diego, M.; Carosi, G.; Dhumeaux, D.; Craxi, A.; Lin, A.;
Hoffman, J.; Yu, J. N. Engl. J. Med 2002, 347, 975.
Mean pharmacokinetic parameters for 26 in rat at 5 mg/kga
IV dose (5 mg/kg)
Oral dose (5 mg/kg)
b
b
t1/2
Vss
1.87
Vb
AUC
7.3
CLp
t1/2
Cmax
Tmax
AUC
6.4
F
4.3
6.11
0.70
4.3
0.58
3.00
94.3
t1/2 (h); Vss (L/kg); Vb (L/kg); AUC0ꢀ1
(l
g h/mL); CLp (L/h kg).
a
6. Simmonds, P. J. Hepatology 1999, 31, 54.
7. Moradpour, D.; Brass, V.; Gosert, R.; Woelk, B.; Blum, H. E. Trends Mol. Med.
2002, 8, 476.
All parameters are means of values obtained from each of three rats.
Harmonic mean.
b
8. Gordon, C. P.; Keller, P. A. J. Med. Chem. 2005, 48, 1.
9. Dhanak, D.; Duffy, K. J.; Johnston, V. K.; Lin-Goerke, J.; Darcy, A. N.; Gu, B.;
Silverman, C.; Gates, A. T.; Nonnemacher, M. R.; Earnshaw, D. L.; Casper, D. J.;
Kaura, A.; Baker, A.; Greenwood, C.; Gutshall, L. L.; Maley, D.; DelVecchio, A.;
Macarron, R.; Hofmann, G. A.; Alnoah, Z.; Cheng, H.-Y.; Chan, G.; Khandekar, S.;
Keenan, R. M.; Sarisky, R. T. J. Biol. Chem. 2002, 277, 38322.
10. (a) Ukrainets, I. V.; Taran, E. A.; Gorokhova, O. V.; Jaradat, N. A.; Voronina, L. N.;
Porokhnyak, I. V. Chem. Heterocycl. Compd. (Engl. Trans.) 2000, 36, 346; (b)
Kovalenko, S. N.; Chernykh, V. P.; Shkarlat, A. I.; Ukrainets, I. V.; Gridasov, V. I.;
Rudnev, S. A. Chem. Heterocyl. Compd. (Engl. Trans.) 1998, 34, 791.
11. Gu, B.; Johnston, V. K.; Gutshall, L. L.; Nguyen, T. T.; Gontarek, R. R.; Darcy, M.
G.; Tedesco, R.; Dhanak, D.; Duffy, K. J.; Kao, C. C.; Sarisky, R. T. J. Biol. Chem.
2003, 278, 16602.
Table 3
Mean liver/plasma levels of 26 after 5 mg/kg oral dose in rata
Concentration (
6
l
g/g or
lg/mL)
Time (h)
1
12
24
0.15
0.0
Liver concentration
Plasma concentration
Liver/plasma ratio
14.2
0.45
34.3
20.8
0.41
48.2
4.6
0.25
17.3
—
a
All parameters are means of values obtained for 26 from each of three rats.
12. Pratt, J. K.; Donner, P.; McDaniel, K. F.; Maring, C. J.; Kati, W. M.; Mo, H.;
Mittleton, T.; Liu, Y.; Ng, T.; Xie, Q.; Zhang, R.; Montgomery, D.; Molla, A.;
Kempf, D. J.; Kohlbrenner, W. Bioorg. Med. Chem. Lett. 2005, 15, 1577.
13. Rosen, T.; Chu, D. T. W.; Lico, I. M.; Fernandes, P. B.; Marsh, K.; Shen, L.; Cepa, V.
G.; Pernet, A. G. J. Med. Chem. 1988, 31, 1598.
14. Buckle, D. R.; Cantello, B. C. C.; Smith, H.; Smith, R. J.; Spicer, B. A. J. Med. Chem.
1977, 20, 1059. Also see: Hutchinson, D.K. et al. U.S. Patent Application US
2005/0107364, May 19, 2005.
15. Laganis, E. D.; Chenard, B. L. Tetrahedron Lett. 1984, 25, 5831.
16. Ward, D. E.; Rhee, C. K. Tetrahedron Lett. 1991, 32, 7165.
17. Rathke, M. W.; Cowan, P. J. J. Org. Chem. 1985, 50, 2622.
18. (a) Barbero, M.; Cadamuro, S.; Digani, I.; Fochi, R.; Gatti, A.; Regondi, V.
Synthesis 1988, 22; (b) Gompper, R.; Kutter, E. Chem. Ber. 1965, 98, 1365.
19. Walter, L. A. J. Am. Chem. Soc. 1943, 65, 738.
methanesulfonamide 26 showed surprisingly good activity both in
enzymatic and cell culture assays. Most notably, 26 showed good
pharmacokinetic behavior in the rat and showed outstanding
selective penetration into the liver. These data taken together sug-
gested that further work on analogs of 26 with unsymmetrical al-
kyl substitution on the B ring would be justified, and these studies
have been ongoing in our laboratories.
Supplementary data
20. Topliss, J. G.; Sherlock, M. H.; Reimann, H.; Konzelman, L. M.; Shapiro,
E. P.; Pettersen, B. W.; Schneider, H.; Sperber, N. J. Med. Chem. 1963,
6, 122.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2008.06.043.
21. Tedesco, R.; Shaw, A. N.; Bambal, R.; Chai, D.; Concha, N. O.; Darcy, M. G.;
Dhanak, D.; Fitch, D. M.; Gates, A.; Gerhardt, W. G.; Halegoua, D. L.; Han, C.;
Hofmann, G. A.; Johnston, V. K.; Kaura, A. C.; Liu, N.; Keenan, R. M.; Lin-Goerke,
J.; Sarisky, R. T.; Wiggall, K. J.; Zimmerman, M. N.; Duffy, K. J. J. Med. Chem.
2006, 49, 971.
References and notes
1. Choo, Q. L.; Kuo, G.; Weiner, A. J.; Overby, L. R.; Bradley, D. W.; Houghton, M.
Science 1989, 244, 359.