Discovery of a novel series of non-nucleoside thumb pocket 2 HCV NS5B polymerase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2013.02.110

A novel series of non-nucleoside thumb pocket 2 HCV NS5B polymerase inhibitors were derived from a fragment-based approach using information from X-ray crystallographic analysis of NS5B-inhibitor complexes and iterative rounds of parallel synthesis. Structure-based drug design strategies led to the discovery of potent sub-micromolar inhibitors 11a-c and 12a-c from a weak-binding fragment-like structure 1 as a starting point.

Full text of DOI:10.1016/j.bmcl.2013.02.110

Accepted Manuscript
Discovery of a Novel Series of Non-Nucleoside Thumb Pocket 2 HCV NS5B
Polymerase Inhibitors
Timothy A. Stammers, René Coulombe, Jean Rancourt, Bounkham
Thavonekham, Gulrez Fazal, Sylvie Goulet, Araz Jakalian, Dominic Wernic,
Youla Tsantrizos, Marc-André Poupart, Michael Bös, Ginette McKercher,
Louise Thauvette, George Kukolj, Pierre L. Beaulieu
PII:
S0960-894X(13)00297-7
DOI:
http://dx.doi.org/10.1016/j.bmcl.2013.02.110
BMCL 20228
Reference:
To appear in:
Bioorganic & Medicinal Chemistry Letters
Received Date:
Accepted Date:
5 February 2013
25 February 2013
Please cite this article as: Stammers, T.A., Coulombe, R., Rancourt, J., Thavonekham, B., Fazal, G., Goulet, S.,
Jakalian, A., Wernic, D., Tsantrizos, Y., Poupart, M-A., Bös, M., McKercher, G., Thauvette, L., Kukolj, G.,
Beaulieu, P.L., Discovery of a Novel Series of Non-Nucleoside Thumb Pocket 2 HCV NS5B Polymerase Inhibitors,
Bioorganic & Medicinal Chemistry Letters (2013), doi: http://dx.doi.org/10.1016/j.bmcl.2013.02.110
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers
we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and
review of the resulting proof before it is published in its final form. Please note that during the production process
errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Graphical Abstract
To create your abstract, type over the instructions in the template box below.
Fonts or abstract dimensions should not be changed or altered.
Discovery of a Novel Series of Non-
Nucleoside Thumb Pocket 2 HCV NS5B
Polymerase Inhibitors
Leave this area blank for abstract info.
Timothy A. Stammersa,*, René Coulombea, Jean Rancourta, Bounkham Thavonekhama, Gulrez Fazala, Sylvie
Gouleta, Araz Jakaliana, Dominic Wernica, Youla Tsantrizosa, Marc-André Pouparta, Michael Bösa, Ginette
McKercherb, Louise Thauvetteb, George Kukoljb and Pierre L. Beaulieua
O
O
Cl
Cl
O
S
O
OH
OH
Cl
O
Parallel Synthesis
Rationale Design
NH
S
O
NH
S
HN
N
O
O
O
1
11b
12b
O
O
IC50 100 M
IC50 0.22 M
IC50 0.16 M

Discovery of a Novel Series of Non-Nucleoside Thumb
Pocket 2 HCV NS5B Polymerase Inhibitors
Timothy A. Stammersa,*, René Coulombea, Jean Rancourta, Bounkham Thavonekhama, Gulrez
Fazala, Sylvie Gouleta, Araz Jakaliana, Dominic Wernica, Youla Tsantrizosa, Marc-André
Pouparta, Michael Bösa, Ginette McKercherb, Louise Thauvetteb, George Kukoljb and Pierre L.
Beaulieua
a Department of Chemistry, Boehringer Ingelheim (Canada) Ltd., Research and Development, 2100 rue Cunard,
Laval (Québec), Canada, H7S 2G5
b Department of Biological Sciences, Boehringer Ingelheim (Canada) Ltd., Research and Development, 2100 rue
Cunard, Laval (Québec), Canada, H7S 2G5
Abstract:
A novel series of non-nucleoside thumb pocket 2 HCV NS5B polymerase inhibitors were derived from a
fragment-based approach using information from x-ray crystallographic analysis of NS5B-inhibitor
complexes and iterative rounds of parallel synthesis. Structure-based drug design strategies led to the
discovery of potent sub-micromolar inhibitors 11a-c and 12a-c from a weak-binding fragment-like
structure 1 as a starting point.
The hepatitis C virus is a leading cause of liver disease and represents a serious health
concern.1 Treatment of chronic HCV infection is based on the poorly tolerated combination of
pegylated interferon-alpha (peg-IFN) and the nucleoside analogue ribavirin (RBV).2 Historically,
this regimen had limited efficacy against the genotype (gt) 1 strains3 that are predominant in
Europe, North America and Japan4 providing sustained virological response (SVR) rates of 40-
50%. In 2011, the first direct acting HCV antivirals, the protease inhibitors telaprevir and
boceprevir, were approved in combination with peg-IFN and RBV for the treatment of HCV gt 1
infection. These new agents improved SVR rates to approximately 68-75% while decreasing
treatment duration for a significant proportion of patients.5 The HCV drug development
landscape is rapidly evolving towards the use of combinations of direct acting antivirals that will
eliminate the need for peg-IFN and provide better efficacy and tolerability for the treatment of
HCV infection.6 We and others have been working on various approaches to find new,
complementary oral antivirals that act directly against virally encoded functions.7
To complement our efforts in the development of thumb pocket 1 non-nucleoside
inhibitors (NNIs) of the HCV NS5B polymerase,8 we began an initiative to identify new lead
structures that bind to one of the other three reported allosteric sites of the enzyme9 and could
be used in combination with our thumb pocket 1 inhibitor BI 2071278b and HCV protease
inhibitor faldaprevir.10 In a search for low molecular weight ligands that bind to the NS5B protein
and could serve as starting points for a fragment-based approach, we discovered that
sulfonamide derivative 1 binds in the previously identified thumb pocket 2 allosteric pocket
(Figure 1).11 It is notable that 1 was also observed in palm site 1. However, it was the
interaction of 1 in thumb pocket 2 that was successfully translated into a novel lead series.
1

Cl
H
N
O
N
S
O
O
Cl
NH
Cl
Cl
O
Cl
O
OH
1
2
Figure 1. Ribbon structure of the HCV NS5B polymerase.
Colored in red is the palm domain that includes the active
site. The finger domain is colored in blue and the thumb
domain is green. Sulfonamide 1 is depicted as a CPK
model bound to thumb pocket 2 (green domain) and palm
site 1 (green/red domain interface).
A pocket view overlay of fragment 1 with previously reported pyranoindole thumb pocket
2 inhibitor 212 (Figure 2A) revealed a good overlap of the halogenated benzene rings over the
lipophilic surface defined by residues L419 and M423 as well as the occupancy of the deep
lipophilic pocket by the methylisoxazole of 1 and the propyl chain of 2. A key interaction that
was observed with the carboxylic acid of pyranoindole 2 but not observed for 1 was a hydrogen
bond with S476.
2

A
B
C
Figure 2. X-ray co-structure overlays in thumb pocket 2
A) Fragment hit 1 (white) and the pyranoindole derivative 2 (orange)
B) Fragment hit 1 (white) and the anthranilic acid derivative 5c (magenta)
C) Anthranilic acid derivatives: Trichlorophenylsulfonamide derivative 5c (magenta) and the 5-
bromothiophenylsulfonamide 7d (green)
In an attempt to incorporate this interaction into fragment 1, 41 anthranilic acid
sulfonamides 5 were synthesized from anthranilic acid derivatives 4 and 2,4,5-trichlorosulfonyl
chloride 3 (Equation 1). Improved potency was observed as summarized by the representative
analogues 5a-c in table 1. For example, unsubstituted anthranilic acid 5a was 4-fold more
active than 1 (IC50~100 M) and halogen substituents at the 4- and 5-positions (compounds 5b
& 5c) provided inhibitors with IC50 values in the 10 μM range.
3

O
H2N
OH
Cl
Cl
Cl
Cl
4
O
R1
a
S
S
O
O
O
Cl
3
Cl
Cl
5
Equation 1. Sulfonylation of anthranilic acid derivatives
a) 4, aniline, sat. aq. Na2CO3, 2 d, RT
Table 1. Carboxylic acid containing inhibitors
R
I C ( M)
50
O
OH
H
N
26
5a
6
5
3
4
O
O
OH
H
N
9. 0
11
5b
5c
Br
OH
H
N
Cl
Analysis of the inhibitor:enzyme complex of 4-chloroanthranilic acid derivative 5c
indicated that this analogue was partially successful in achieving the desired hydrogen bonding
interaction. Figure 2B shows the overlay with fragment 1 where the carboxylic acid of 5c forms
a water-mediated H-bond with the backbone amide N-H of S476.
The SAR of the arylsulfonamide substituent was explored in combination with the 5-
bromo- and 4-chloroanthranilic acid substitution of 5b-c. Halogenated anthranilic acids 4b-c
were coupled with arylsulfonyl chlorides under the same reaction conditions described in
Equation 1 to produce compound series 6 (n=74) and 7 (n=54). Unfortunately, no significant
improvements in potency were achieved. Representative inhibitors 6a-d and 7c-d are shown in
Table 2. The unsubstituted phenyl sulfonamide 6a was approximately 4-fold less potent than
2,4,5-trichlorosulfonylamide 5b. Halogenated aryl- and heteroarylsulfonamides such as 6b-d
and 7b-d resulted in a relatively flat SAR. Despite providing only a marginal improvement over
4

5c, the most important inhibitor to emerge from this exercise was 5-bromothien-2-ylsulfonamide
7d. Analysis of the binding mode of this compound provided a key observation that led us
towards inhibitors with improved potency.
Table 2. 5-Bromo- and 4-chloroanthranilic acid sulfonamides
OH
OH
O
O
H
H
R2
N
R2
N
S
S
O
O
O
O
Br
Cl
6
7
R2
IC50 (M)
IC50 (M)
6a
6b
43
Cl
Cl
8.4
F
Br
6c
6d
7c
7d
6.8
4.6
4.5
6.1
F
Br
S
Analysis of the polymerase:7d complex highlighted an unexpected change in binding
mode as shown in Figure 2C. In the overlay of 7d and 5c, the anthranilic acid and sulfonamide
moieties were transposed to enable bidentate hydrogen bonding between the inhibitor
carboxylic acid and the backbone N-H’s of S476 and Y477. These hydrogen bonding patterns
in thumb pocket 2 are similar to those reported for the thiophene carboxylic acid inhibitor 8.11f
O
S
OH
N
O
8
The new binding mode prompted an exploration of the 4- and 5- positions of the
anthranilic acid ring in order to improve potency by exploiting hydrophobic contacts with the
protein surface in a similar fashion to the halogenated phenyl rings of 2 and 5c in figure 2A and
2B. As shown in Table 3, introduction of a phenyl group to the anthranilic acid scaffold,
analogous to thiophene carboxylic acid 8, at either the 5- (R5=Ph, 9) or 4- (R4=Ph, 10) position
5

did not improve potency. After examining a variety of substituents at these positions,
incorporation of an oxygen linker between the two phenyl rings was found to provide a 10-fold
improvement in potency. This improvement was observed for both the 4-phenoxy 11a and 5-
phenoxy 12a derivatives.
Table 3. Anthranilic acid substitution at the 5- and 4- positions
O
5
R5
OH
R4
NH
S
4
O
S
Br
O
IC50 (M)
R4
R5
9
3.7
H
H
10
6.7
O
11a
12a
0.55
0.61
H
H
O
Scheme 1 describes the introduction of phenyl substitution at the 4- and 5- positions of
the anthranilic acid scaffold. Phenylboronic acid was coupled to 5-bromoanthranilic acid methyl
ester 13 or 4-bromo-2-nitrobenzoic acid 14 using a Suzuki-Miyaura coupling to produce the
corresponding intermediates 15 and 16. The 5-phenylanthranilic acid methyl ester 15 was
reacted with 5-bromo-2-thiophenesulfonyl chloride then saponified to afford inhibitor 9. Methyl-
2-nitro-4-phenylbenzoate 16 was reduced with Pd(OH)2/C prior to sulfonylation and
saponification to prepare inhibitor 10. The synthesis of the 4- and 5-phenoxy substituted
anthranilic acids 11a-c and 12a-c commenced with the SNAr addition of phenol on methyl-5-
chloro-2-nitrobenzoate 17 or methyl-4-fluoro-2-nitrobenzoic ester 18. The nitro groups were
reduced by hydrogenolysis followed by treatment with HCl to produce aniline salts 19 and 20.
Sulfonylation and saponification afforded 4- and 5-phenoxyanthranilic acid inhibitors 11a-c and
11a-c.
6

O
O
Br
O
O
9
b, c
a
d
NH2
O
NH2
O
13
15
OH
+ O
O
10
+ O
e, b, c
Br
Cl
N
O
N
O
O
14
17
16
19
O
O
O
O
11a-c
12a-c
f, g
b, c
+ O
N
NH3Cl
O
O
O
O
O
+ O
f, g
b, c
F
N
O
O
NH3Cl
18
20
Scheme 1. Synthesis of phenyl- and phenoxyanthranilic
acid analogues 9-12.
(a) Phenylboronic acid (1.2 eq.), (Ph3P)4Pd (0.2 eq.), 2 M
aq. Na2CO3 (2 eq.), DME, 80 oC, 20 h, 57%; (b) sulfonyl
chloride (3 eq.), pyridine, 50 oC, 8 h, 80%; (c) NaOH (5
eq.), DMSO, RT, 0.5 h, 60%; (d) same as (a) followed
treatment of the crude product with CH2N2 in ether prior to
purification, 88%; (e) Pd(OH)2/C (0.2 eq.), MeOH, 1 atm
H2, 16 h, 96%; (f) Phenol (1.5 eq.), K2CO3 (1.5 eq.),
DMSO, 90 oC, 6 h, 57%; (g) same as (e) follow by filtration
and treatment with HCl in ether, 79% 2 steps.
The SAR of sulfonamide substituents on the phenoxyanthranilic acid inhibitors was
similar to that reported for the thiophene carboxylic acid chemotype (e.g. 8).13 Parallel SAR was
observed for the 4- and 5-phenoxyanthranilic acid isomers. The 4-methylphenylsulfonamides-
11b/12b and the 4-bromo-2-fluorophenylsulfonamides 11c/12c (Table 4) were the most potent
inhibitors identified from a combined total of 47 inhibitors prepared with the 4- and 5-
phenoxyanthranilic acids.
Table 4. 5- and 4-phenoxy anthranilic acid sulfonamide inhibitors
O
O
O
OH
OH
NH
S
O
NH
S
O
O
R
R
O
O
R
IC50 (M)
IC50 (M)
11b
11c
0.22
0.16
12b
12c
0.34
0.31
F
Br
7

We studied the binding mode of the 4-phenoxy- (11b) and 5-phenoxy- (12b) anthranilic
acid inhibitors in thumb pocket 2. The binding orientation of the anthranilic acid scaffold
remains consistent with 7d as can be seen by comparing Figure 2C with the overlay of 11b/12b
in Figure 3. However, the phenoxy substituent of both isomers induced an important change in
the pocket conformation. A coordinated movement of residues M423 and L497 created a
shallow pocket for the phenoxy group while concomitantly expanding the deep hydrophobic
pocket to accommodate larger substituents such as the 2-fluoro-4-bromophenyl group of 11c
and 12c.11h The observation that the phenoxy group occupied the same shallow lipophilic
pocket whether they were projected from the 5- (11b) or 4- (12b) position of the anthranilic acid
also provides a rationale for the comparable potency.
Figure 3. X-ray co-structure overlay of 5-
phenoxy (11b, orange) and 4-phenoxy
(12b, green) anthranilic acid inhibitors
bound in the induced conformation of
thumb pocket 2.
Three compounds (11b, 12b and 11c) were profiled against a panel of thumb pocket
1,14, 15 thumb pocket 216, 17 and palm site 118 single amino acid site-specific mutants of the HCV
NS5B polymerase and the results are shown in table 5. Consistent with binding in thumb
pocket 2, the potency of each compound shifted approximately 15-fold against the previously
disclosed L419M mutant.16 Minimal changes in sensitivity were observed with the M423T
thumb pocket 2 mutant. This is notable as mutations at M423 have been predominately
selected in HCV infected patients dosed with thumb pocket 2 inhibitors.19, 20 As expected, no
shift was observed between the wild type genotype 1b and thumb pocket 1 (P495S) or palm site
1 (M414T) mutants. High selectivity was observed versus the RNA-dependent RNA-
polymerase from poliovirus and the mammalian RNA polymerase II isolated from calf thymus.21
Unfortunately, inhibitors 11b, 12b and 11c were not active in the HCV cell-based replicon
assay.
Table 5. Biochemical profiling of selected phenoxyanthranilic acid sulfonamide inhibitors..
Wild type Thumb 2 Thumb 2
Thumb 1 Palm 1 Polio virus Calf thymus
P495S M414T polymerase polymerase
IC50, M IC50, M IC50, M
Cmpds
NS5B
IC50, M
0.086
M423T
IC50, M
0.17
L419M
IC50, M
1.2
IC50, M
>100
11b
12b
11c
--
0.077
0.093
--
0.052
0.052
>100
>100
>100
0.072
0.11
0.10
0.20
1.2
1.8
>100
>100
8

The available in vitro ADME profiles of representative compounds 11b, 12b and 11c are
summarized in Table 6. Moderate to poor Caco-2 permeability may be a reflection of poor
membrane permeability and would be consistent with the lack of cell culture activity of these
compounds. Poor stability in the presence of human liver microsome and low micromolar
CYP450 inhibition against select isozymes will need to be addressed during the optimization of
the chemical series. By virtue of the presence of a carboxylic acid, excellent solubility was
observed at neutral pH.
Table 6. Summary of in vitro ADME properties of selected anthranilic acid sulfonamide inhibitors.
Solubilitya Caco-2b
HLMc
CYP450 IC50
Cmpds
11b
12b
(μg/mL) (x10-6 cm/s) (t1/2, min.) 1A2, 2C9, 2C19, 2D6, 3A4
>900
>700
>900
2.2
2.4
<0.1
11
17
27
1.8, 3.0, >30, >30, >30
>30, 3.1, 7.9, >30, >30
>30, 1.1, 1.1, >30, 26
11c
a Determined on amorphous solids, 24 hour solubility in pH 7.2 buffer using the shaking flask method.
b Apical to Basal permeability at pH 7.4.
c Half-life measured in the presence of human liver microsomes at an initial concentration of 10 M.
We have used multiple iterations of parallel synthesis and structure based drug design to
discover a novel lead series of thumb pocket 2 anthranilic acid inhibitors from a low affinity
fragment. A 650-fold improvement in biochemical potency was achieved for compound 12b
from fragment 1. Additional profiling confirmed that these inhibitors are selective NS5B thumb
pocket 2 inhibitors. Efforts directed towards establishing cell culture activity and improving in
vitro ADME properties for this class of allosteric HCV polymerase inhibitors will be reported in
due course.
Acknowledgements
We thank Serge Valois, Colette Boucher, Sylvain Bordeleau and Michael Little for analytical
support as well as Josie DeMarte and Manon Rhéaume for ADME profiling.
References and notes
1
Lavanchy, D. Clin. MicroBiol. Infect. 2011, 17, 107.
Ghany, M. G.; Strader, D. B.; Thomas, D. L.; Seeff, L. B Hepatology 2009, 49, 1335.
Farnik, H.; Mihm, U.; Zeuzem, S. Liver Int. 2009, 29(s1), 23.
Who Health Organization Initiative for Vaccine Research, Viral Cancers, Hepatitis C Virus:
2
3
4
http://www.who.int/vaccine_research/diseases/viral_cancers/en/index2.html
5
Marks, K. M.; Jacobson, I. Antiviral Therapy 2012, 17, 1119.
(a) Gane, E. Antiviral Therapy 2012, 17, 1210. (b) Pockros, P. J. Drugs 2012, 72, 1825.
(a) Doyle, J. S.; Aspinall, E.; Liew, D.; Thompson, A. J.; Hellard, M. E. Br. J. Clin. Pharm. 2012, ASAP.
6
7
(b) Lange, C. M.; Zeuzem, S. J. Hepatology 2012, ASAP.
8 (a) Beaulieu P. L.; Bös, M.; Cordingley, M. G.; Chabot, C. ; Fazal, G.; Garneau, M.; Gillard, J. R.;
Jolicoeur, E.; LaPlante, S.; McKercher, G.; Poirier, M. ; Poupart, M-A ; Tsantrizos, Y. S. ; Duan, J.;
Kukolj, G. J. Med. Chem. 2012, 55, 7650. (b) Larrey, D.; Lohse, A. W.; de Ledinghen, V.; Trepo, C.;
Gerlach, T.; Zarski, J.-P.; Tran, A.; Mathurin, P.; Thimme, R.; Arastéh, K.; Trautwein, C.; Cerny, A.;
Dikopoulos, N.; Schuchmann, M.; Heim, M. H.; Gerken, G.; Stern, J. O.; Wu, K.; Abdallah, N.; Girlich, B.;
Scherer, J.; Berger, F.; Marquis, M.; Kukolj, G.; Böcher, W.; Steffgen, J. J. Hepatol. 2012, 57, 39. (c)
Beaulieu P. L.; Chabot, C.; Duan, J.; Garneau, M.; Gillard, J.; Jolicoeur, E.; Poirier, M.; Poupart, M.-A.;
Stammers, T.A.; Kukolj, G.; Tsantrizos, Y. S. Bioorg. Med. Chem. Lett. 2011, 21, 3664. (d) Beaulieu P. L.;
Gillard, J.; Jolicoeur, E.; Duan, J.; Garneau, M.; Kukolj, G.; Poupart, M.-A. Bioorg. Med. Chem. Lett.
2011, 21, 3658. (e) Beaulieu P. L.; Dansereau, N.; Duan, J.; Garneau, M.; Gillard, J.; McKercher, G.;
LaPlante, S. ; Lagacé, L.; Thauvette, L.; Kukolj, G. Bioorg. Med. Chem. Lett. 2010, 20, 1825. (f) Beaulieu
P. L.; Jolicoeur, E.; Gillard, J.; Brochu, C.; Coulombe, R.; Dansereau, N.; Duan, J.; Garneau, M.; Jakalian,
A.; Kühn, P.; Lagacé, L.; LaPlante, S. ; McKercher, G.; Perreault, S.; Poirier, M.; Poupart, M.-A.;
9

Stammers, T.A.; Thauvette, L.; Thavonekham, B.; Kukolj, G. Bioorg. Med. Chem. Lett. 2010, 20, 857. (g)
Goulet, S.; Poupart, M.-A.; Poirier, M.; Gillard, J.; Kukolj, G.; Beaulieu P. L.; Bioorg. Med. Chem. Lett.
2010, 20, 196 and references therein.
9 For recent review on the different classes of HCV NS5B inhibitors see: (a) Sofia, M. J.; Chang, W.;
Furman, P. A.; Mosley, R. T.; Ross, B. S. J. Med. Chem. 2012, 55, 2481. (b) Li, H.; Shi, S. T. Future Med
Chem. 2010, 2, 121. (c) Legrand-Abravanel, F.; Nicot, F.; Izopet, J. Expert Opin. Investig. Drugs 2010,
19, 963. (d) Beaulieu, P.L. Expert Opin. Ther. Patents 2009, 19, 145.
10 (a) Llinás-Brunet, M.; Bailey, M. D.; Goudreau, N.; Bhardwaj, P. K.; Bordeleau, J.; Bös, M.; Bousquet,
Y.; White, P. W.; Cordingley, M. G.; Duan, J.; Forgione, P.; Garneau, M.; Ghiro, E.; Gorys, V.; Goulet, S.;
Halmos, T.; Kawai, S. H.; Naud, J.; Poupart, M.-A.; White, P. W.; J. Med. Chem. 2010, 53, 6466. (b)
White, P. W.; Llinas-Brunet, M.; Amad, M.; Bethell, R. C.; Bolger, G.; Cordingley, M. G.; Duan, J.;
Garneau, M.; Lagacé, L.; Thibeault, D.; Kukolj, G. Antimicrobial Agents and Chemotherapy 2010, 54,
4611. (c) Lemke, C. T.; Goudreau, N.; Zhao, S.; Hucke, O.; Thibeault, D.; Llinás-Brunet, M.; White, P. W.;
J. Biol. Chem. 2011, 286, 11434 and references therein.
11 References containing biochemical characterization and x-ray co-structures of representative
chemotypes that bind to thumb site 2: (a) Kumar, D. V.; Rai, R.; Brameld, K. A.; Somoza, J. R.;
Rajagopalan, R.; Janc, J. W.; Xia, Y. M.; Ton, T. L.; Shaghafi, M. B.; Hu, H.; Lehoux, I.; To, N.; Young, W.
B.; Green, M. J. Bioorg. Med. Chem. Lett. 2011, 21, 82. (b) Ontoria, J. M.; Rydberg, E.; H.; Di Marco, S.;
Tomei, L.; Attenni, B.; Malancona, S.; Hernando, J. I. M.; Gennari, N.; Koch, U.; Narjes, F.; Rowley, M.;
Summa, V.; Carroll, S. S.; Olsen, D.; De Francesco, R.; Altamura, S.; Migliaccio, G.; Carfi, A. J. Med.
Chem. 2009, 52, 5217. (c) Antonysamy, S. S.; Aubol, B.; Blaney, J.; Browner, M. F.; Giannetti, A. M.;
Harris, S. F.; Hébert, N.; Hendle, J.; Hopkins, S.; Jefferson, E.; Kissinger, C.; Leveque, V.; Marciano, D.;
McGee, E.; Nájera, I.; Nolan, B.; Tomimoto, M.; Torres, E.; Wright, T. Bioorg. Med. Chem. Lett. 2008, 18,
2990. (d) Yan, S.; Appleby, T.; Larson, G.; Wu, J. Z.; Hamatake, R K.; Hong, Z.; Yao, N. Bioorg. Med.
Chem. Lett. 2007, 17, 1991. (e) Howe, A. Y. M.; Cheng, H.; Thompson, I.; Chunduru, S. K.; Herrmann, S.;
O’Connell J.; Agarwal, A.; Chopra, R.; Del Vecchio, A. M. Antimicrob. Agents and Chemother. 2006, 50,
4103. (f) Biswal, B. K.; Cherney, M. M.; Wang, M.; Chan, L.; Yannopoulos, C. G.; Bilimoria, D.; Nicolas,
O.; Bedard, J.; James, M. N. G. J. Biol. Chem. 2005, 280, 18202. (g) Wang, M.; Ng, K. K.-S.; Cherney, M.
M.; Chan, L.; Yannopoulos, C. G.; Bedard, J.; Morin, N.; Nguyen-Ba, N.; Alaoui-Ismaili, M. H.; Bethell, R.
C.; James, M. N. G. J. Biol. Chem. 2003, 278, 9489. (h) Love, R. A.; Parge, H. E.; Yu, X.; Hickey, M. J.;
Diehl, W,; Gao, J.; Wriggers, H.; Ekker, A.; Wang, L.; Thomson, J. A.; Dragovich, P. S.; Fuhrman, S. A. J.
Virol. 2003, 77, 7575.
12 Gopalsamy, A.; Lim, K.; Ciszewski, G.; Park, K.; Ellingboe, J. W.; Bloom, J.; Insaf, S.; Upeslacis, J.;
Mansour, T. S.; Krishnamurthy, G.; Damarla, M.; Pyatski, Y.; Ho, D.; Howe, A. Y. M.; Orlowski, M.; Feld,
B.; O’Connell, J. J. Med. Chem. 2004, 47, 6603.
13 Chan, L.; Das, S. K.; Reddy, T. J.; Poisson, C.; Proulx, M.; Pereira, O.; Courchesne, M.; Roy, C.;
Wang, W.; Siddiqui, A.; Yannopoulos, C. G.; Nguyen-Ba, N.; Labrecque, D.; Bethell, R.; Hamel, M.;
Courtmanche-Asselin, P.; L’Heureux, L.; David, M.; Nicolas, O.; Brunette, S.; Bilimoria, D.; Bédard, J.
Bioorg. & Med. Chem. Lett. 2004, 14, 793.
14 Tomei, L.; Altamura, S.; Bartholomew, L.; Biroccio, A.; Ceccacci, A.; Pacini, L.; Narjes, F.; Gennari, N.;
Biscocci, M.; Incitti, I.; Orsatti, L.; Harper, S.; Stansfield, I.; Rowley, M.; De Francesco, R.; Migliaccio, G.
J. Virol. 2003, 77, 13225.
15 Kukolj, G.; McGibbon, G. A.; McKercher, G.; Marquis, M.; Lefebvre, S.; Thauvette, L.; Gauthier, J.;
Goulet, S.; Poupart, M.-A.; Beaulieu, P. L. J. Biol. Chem. 2005, 280, 39260.
16 Le Pogam, S.; Kang, H.; Harris, S. F.; Leveque, V.; Giannetti, A. M.; Ali, S.; Jiang, W.-R.; Rajyaguru,
S.; Tavares, G.; Oshiro, C.; Hendricks, T.; Klumpp, K.; Symons, J.; J.; Browner, M. F.; Cammack, N.;
Najera, I.; J. Virol. 2006, 80, 6146.
17 Shi, S. T.; Herlihy, K. J.; Graham, J. P.; Nonomiya, J.; Rahavendran, S. V.; Skor, H.; Irvine, R.; Binford,
S.; Tatlock, J.; Li, H.; Gonzalez, J.; Linton, A.; Patrick, A. K.; Lewis, C. Antimicrob. Agents and
Chemother. 2009, 53, 2544.
18 Nguyen, T. T.; Gates, A. T.; Gutshall, L. L.; Johnston, V. K.; Gu, B.; Duffy, K. J.; Sarisky, R. T.
Antimicrob. Agents and Chemother. 2003, 47, 3525.
19 Troke, P. J. F.; Lewis, M.; Simpson, P.; Gore, K.; Hammond, J.; Craig, C.; Westby, M. Antimicrob.
Agents and Chemother. 2012, 56, 1331.
10

20 Cooper, C.; Lawitz, E. J.; Ghali, P.; Rodriguez-Torres, M.; Anderson, F. H.; Lee, S. S.; Bédard, J.;
Chauret, N.; Thibert, R.; Boivin, I.; Nicolas, O.; Proulx, L. J. Hepatol. 2009, 51, 39.
21 McKercher, G.; Beaulieu, P. L.; Lamarre, D.; LaPlante, S., Lefebvre, S.; Pellerin, C.; Thauvette, L.;
Kukolj, G. Nucleic Acids Research 2004, 32, 422.
11