High potency improvements to weak aryl uracil HCV polymerase inhibitor leads

Article abstract of DOI:10.1016/j.bmcl.2013.05.078

Described herein is the development of a potent non-nucleoside, small molecule inhibitor of genotype 1 HCV NS5B Polymerase. A 23 μM inhibitor that was active against HCV polymerase was further elaborated into a potent single-digit nanomolar inhibitor of HCV NS5B polymerase by additional manipulation of the R and R₁ substituents. Subsequent modifications to improve physical properties were made in an attempt to achieve an acceptable pharmacokinetic profile.

Full text of DOI:10.1016/j.bmcl.2013.05.078

Bioorganic & Medicinal Chemistry Letters 23 (2013) 4367–4369
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
High potency improvements to weak aryl uracil HCV polymerase
inhibitor leads
⇑
Pamela Donner , John T. Randolph, Peggy Huang, Rolf Wagner, Clarence Maring, Ben Hock Lim,
Lynn Colletti, Yaya Liu, Rubina Mondal, Jill Beyer, Gennadiy Koev, Kennan Marsh, David Beno,
Kenton Longenecker, Tami Pilot-Matias, Warren Kati, Akhter Molla, Dale Kempf
AbbVie, Department R4AJ, Building AP52N, 1 North Waukegan Road, North Chicago, IL 60064, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
Described herein is the development of a potent non-nucleoside, small molecule inhibitor of genotype 1
Received 28 January 2013
Revised 20 May 2013
Accepted 22 May 2013
Available online 4 June 2013
HCV NS5B Polymerase. A 23 lM inhibitor that was active against HCV polymerase was further elaborated
into a potent single-digit nanomolar inhibitor of HCV NS5B polymerase by additional manipulation of the
R and R1 substituents. Subsequent modifications to improve physical properties were made in an attempt
to achieve an acceptable pharmacokinetic profile.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
HCV
NS5B polymerase
Aryl-uracil
Hepatitis C virus infects over 3% of the world population.1 HCV
infection is a leading cause of chronic liver disease and the leading
cause of death from liver disease in the United States.2 The death
rate from HCV is expected to continue to rise through 2015.3 The
goal in treating patients infected with HCV is eradication of the
infection. Recently two new protease inhibitor drugs, boceprevir
and telaprevir, were approved by the FDA. However these drugs
must still be used in combination with interferon, which imparts
several drug-related toxicities. Thus, there is still an urgent need
for new HCV drugs with diverse modes of action, so that they
can eventually be combined and replace the use of interferon in
the treatment of this infection.4
R1
O
H
N
H
N
H
N
O
O
O
O
O
O
HN
N
N
N
O
O
R
OH
1
2
3
Figure 1.
substantial alterations to the core were required. Towards that
end, we utilized commercially available ester 4, to synthesize
substituted amides 3 at R1 (Scheme 1).
A soluble 23 lM fragment 1 (Fig. 1) demonstrated acceptable
in vitro permeability, and consistent with the observed physical
properties, demonstrated plasma exposures in rats after oral dosing
(Table 2). Biochemical studies were done using mutant polymer-
ases that showed this class of molecules were over 100-fold resis-
tant to C316Y, 20-fold resistant to M414T and 40-fold resistant to
Y448H which indicated they bind in the NNI III (palm) region.5
A clear shortcoming of 1 as a lead was its weak activity against
HCV polymerase. An initial goal, therefore, was improvement of
the potency of this compound against the enzyme. Seeking to
ascertain the necessity of the acidic phenol, methylation of the
same resulted in 2. Having established this modification as provid-
ing only a modest improvement in activity (Table 1), more
Starting with the commercially available ester 4, we ipso-
nitrated para to the phenol, followed by optional methylation of
the phenol to provide nitro-benzoate 5. Reduction of the nitro
group gave 6 which was followed by alkylation with acrylic acid
to produce 7. Acylation with urea in acetic acid provided 8, which
was cyclized to the dihydrouracil ester 9. Hydrolysis of 9 and treat-
ment with thionyl chloride provided the central intermediate 10
for the SAR studies. Amides 3 were synthesized by heating the acid
chloride 10 with the appropriate amine to produce the final
compounds.
Phenyl amides, 11a and 11b, showed improved activity relative
to the initial leads 1 and 2. Amide 11a demonstrated a 16-fold
improvement against the genotype 1a isoform of the enzyme and
a dramatic 100-fold improvement against the 1b isoform (Table 1).
While the fold difference of 11b against lead 2 was not as dramatic,
⇑
Corresponding author. Tel.: +1 847 937 3245; fax: +1 847 938 2756.
E-mail address: pamela.l.donner@abbvie.com (P. Donner).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.05.078

4368
P. Donner et al. / Bioorg. Med. Chem. Lett. 23 (2013) 4367–4369
Table 1
Structure–activity relationships
R1
O
H
O
O
N
HN
N
HCV polymerase inhibition
HCV replicon inhibition
O
R
R1
R
1a IC50
(
lM)
1b IC50
(
lM)
1a EC50
(l
M)
1b EC50 (lM)
1
2
NA
NA
H
Me
71
26
23
6.4
>100?
>100?
>100?
>100?
11a
11b
H
Me
4.28
0.89
0.23
0.75
18.6
3.67
6.31
1.03
12
13
H
8.59
2.06
0.94
1.24
14.2
>50
15.3
4.24
NH2
Me
H
N
14
Me
3.42
5.12
NA
NA
O
H
15a
15b
H
Me
0.02
0.01
0.002
0.02
0.39
0.06
0.05
0.02
N
S
O O
16a
16b
H
Me
0.18
0.67
0.03
1.04
4.65
3.40
1.13
0.86
O
S
O O
17a
17b
H
Me
0.46
0.77
0.03
0.83
3.55
5.49
1.04
0.56
N
S
O O
O
S
O
18
H
0.36
0.05
27.8
7.56
N
H
19a
19b
H
Me
0.038
0.02
0.01
0.03
0.59
0.27
0.094
0.05
H
N
S
H
N
O
O
O
O
HN
O
N
O
R
20a
20b
H
Me
0.05
0.11
0.007
0.31
1.96
5.31
0.42
0.58
O
H
N
N
S
O O
IC50 values were determined in a biochemical assay for recombinant HCV genotype 1a and 1b polymerases (obtain reference).
EC50 values in HCV genotype 1a and 1b replicon cells were generated in assays containing 5% fetal calf serum (obtain reference).
Table 2
Pharmacokinetics of HCV polymerase inhibitors
Bioavailability (%)
IV t1/2 (h)
AUC (lg h/mL)
Oral AUC (
l
g h/mL)
Solubility (lM)
Permeability (10eÀ6 cm/s)
2
45
15
1.5
NA
0
3.8
0.5
1.1
NA
0.6
12.36
2.19
3.38
NA
5.57
0.33
0.05
NA
73
12
2
13
74
5.5
1.4
0.5
0.2
0.1
11b
15b
19b
20b
2.54
0
a further improvement in absolute potency against both isoforms
of the enzyme was achieved. Though weak, it is noteworthy that
activity against the HCV Replicon in Huh7 cells is now detectable.
While the benzyl homolog 12 also showed an improvement in
activity relative to 1, the slightly weaker performance in most of
the assays relative to 11a directed us to perform further studies
on the phenyl amide base structure.
Synthesis of the para aniline 13, weakened enzyme inhibition,
as did attenuating the basicity of the aniline by acylation (14).
Notably, sulfonylating the aniline to provide 15a increased the
potency substantially across all assays. Indeed, corresponding
analog 15b further improved activity in the 1a genotype assays.
As such, the synthesis of 15b achieved a potency increase over 1
of 7000-fold and 1100-fold in the genotype 1a and 1b enzyme

P. Donner et al. / Bioorg. Med. Chem. Lett. 23 (2013) 4367–4369
4369
O
O
O
H2N
O2N
O
O
O
c
a,b
O
R
O
OH
6
4
5
d
H
N
O
OH
O
O
O
O
H2N
O
H
O
N
N
O
O
N
f
e
O
O
R
O
R
HO
O
R
O
7
9
8
g
H
H
H
N
R1
O
O
N
O
O
N
O
O
O
O
HN
O
N
N
N
h
OH
Cl
i
O
R
O
R
O
R
3
10
Scheme 1. (a) AcOH, HNO3; (b) MeI, base; (c) RaNi; (d) acrylic acid, heat, toluene; (e) urea, AcOH, heat; (f) concd HCl, heat; (g) 1 M NaOH; (h) SOCl2, heat; (i) substituted
aniline or amine, dioxane, heat.
assays respectively. It was also especially gratifying to observe the
20–60 nM activities against the 1a and 1b HCV replicons. The con-
tribution of this methanesulfonyl group was further investigated
by synthesizing several additional analogs, including mesylates,
16a and 16b, the N-methyl-substituted methyl sulfonamides, 17a
and 17b, and meta regioisomer 18. All of these analogs proved less
potent than the methylsulfonamides 15a and 15b. All these
successful. While the most promising analogs achieved antiviral
potencies in the biochemical and replicon assays of less than
50 nM, they unfortunately manifested physical properties that re-
sulted in suboptimal PK profiles. Addition of a solubilizing group
with retention of acceptable antiviral activity, unfortunately, was
insufficient to address these shortcomings. Therefore, due to the
PK shortcomings of this series, more extensive SAR was not done.
Although the methylsulfonamide moiety contributed to the antivi-
ral activity of these compounds, it introduced sufficient polar sur-
face area to the compounds to render them impermeable. Future
efforts will be directed to addressing the permeability of these
compounds including investigation of SAR surrounding other parts
of the molecule and alternative linking strategies.
analogs had TD50 values of >50
good therapeutic indices.
lM, indicating they could achieve
During these investigations, it was observed that the dihydrour-
acil moiety was subject to minor, but measurable hydrolysis. In an
attempt to address this issue, we synthesized the uracil analogs
19a and 19b. While these modifications improved the stability of
the subsequent molecules, they also produced compounds that re-
tained potent activities similar to 15a and 15b.
Disclosures: This research was sponsored by Abbvie. Abbvie con-
tributed to the research, interpretation of data, writing, reviewing,
and approving the publication.
In vivo characterization of several compounds from Table 1
eventually revealed additional hurdles to our drug discovery pro-
gram. While it was determined that the solubility, permeability,
and oral bioavailability of lead fragment 2 in rat was acceptable
(Table 2), the more highly elaborated analogs performed less well
in rat PK experiments. Oral bioavailability of phenyl amide 11b suf-
fered by experiencing moderate fold reductions in oral bioavail-
ability. However, our most potent analogs tested in rat PK (15b
and 19b) provided almost no plasma exposures of compound fol-
lowing oral administration. Relative to 2, all of these compounds
were less permeable and less soluble in vitro. An attempt was
made to improve the solubility of subsequent analogs with 20a
and 20b being examples of compounds that showed similar solu-
bilities to 2, but with much improved antiviral activity (Tables 1).
Unfortunately, rat PK experiments showed no improvement with
respect to oral exposure and, it was therefore determined that
the sulfonamide group that contributed to antiviral activity was
also the functionality that dramatically impacted permeability
and resulted in the poor oral PK performance of the series.
Donner, Randolph, Huang, Wagner, Maring, Lim, Colletti, Liu,
Beyer, Koev, Marsh, Longenecker, Pilot-Matias, Molla, and Kempf
are all employees of Abbvie. Beno and Mondal are employees of
Abbott.
Acknowledgements
The authors would like to thank Doug Hutchinson, Todd
Rockway, and Michael Tufano for their contributions (all of Abbvie
at the time of their contributions).
References and notes
1. WHO, Hepatitis C. Fact Sheet No. 164. Revised October 2000.
2. Kim, W. R. Hepatology 2002, 36, 30.
3. Pol, S.; Mallet, V. O. Expert Opin. Biol. Ther. 2006, 6, 923.
4. Rice, C. Nature 2011, 474, S8.
5. Liu, Y.; Lim, B. H.; Jiang, W. W.; Flentge, C. A.; Hutchinson, D. K.; Madigan, D. L.;
Randolph, J. T.; Wagner, R.; Maring, C. J.; Kati, W. M.; Molla, A. Bioorg. Med. Chem.
Lett. 2012, 22, 3747.
In summary, our efforts in improving the potency of a 23
lM
fragment toward highly potent HCV NS5B inhibitors was