Design and synthesis of pyridone inhibitors of non-nucleoside reverse transcriptase

Article abstract of DOI:10.1016/j.bmcl.2011.10.027

Next generation NNRTIs are sought which possess both broad spectrum antiviral activity against key mutant strains and a high genetic barrier to the selection of new mutant viral strains. Pyridones were evaluated as an acyclic conformational constraint to

Full text of DOI:10.1016/j.bmcl.2011.10.027

Bioorganic & Medicinal Chemistry Letters 21 (2011) 7344–7350
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of pyridone inhibitors of non-nucleoside
reverse transcriptase
Robert Gomez a, , Samson Jolly a, Theresa Williams a, Thomas Tucker a, Robert Tynebor a, Joe Vacca a,
⇑
Georgia McGaughey b, Ming-Tain Lai c, Peter Felock d, Vandna Munshi c, Daniel DeStefano d,
Sinoeun Touch c, Mike Miller d, Youwei Yan e, Rosa Sanchez f, Yuexia Liang f, Brenda Paton f,
Bang-Lin Wan f, Neville Anthony g
a Department of West Point Discovery Chemistry, Merck Research Labs., 770 Sumneytown Pike, PO Box 4, West Point, PA 19486-0004, United States
b Department of Chemistry Modeling and Informatics, Merck Research Labs., 770 Sumneytown Pike, PO Box 4, West Point, PA 19486-0004, United States
c Department of In Vitro Pharmacology, Merck Research Labs., 770 Sumneytown Pike, PO Box 4, West Point, PA 19486-0004, United States
d Department of ID Antiviral HIV Discovery, Merck Research Labs., 770 Sumneytown Pike, PO Box 4, West Point, PA 19486-0004, United States
e Department of Structural Chemistry, Merck Research Labs., 770 Sumneytown Pike, PO Box 4, West Point, PA 19486-0004, United States
f Department of DMPK Preclinical WP, Merck Research Labs., 770 Sumneytown Pike, PO Box 4, West Point, PA 19486-0004, United States
g Department of Boston Discovery Chemistry, Merck Research Labs., 33 Avenue Louis Pasteur, Boston, MA 02115-5727, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
Next generation NNRTIs are sought which possess both broad spectrum antiviral activity against key
mutant strains and a high genetic barrier to the selection of new mutant viral strains. Pyridones were
evaluated as an acyclic conformational constraint to replace the aryl ether core of MK-4965 (1) and
the more rigid indazole constraint of MK-6186 (2). The resulting pyridone compounds are potent inhib-
itors of HIV RT and have antiviral activity in cell culture that is superior to other next generation NNRTI’s.
Ó 2011 Elsevier Ltd. All rights reserved.
Received 9 August 2011
Revised 5 October 2011
Accepted 7 October 2011
Available online 17 October 2011
Keywords:
Non-nucleoside reverse transcriptase
inhibitor
Pyridone
Pyridinone
HIV
HIV (human immunodeficiency virus) has a high rate of muta-
tion due to its infidelity during replication. As a result, single drug
therapy is ineffective due to the rapid selection of resistant mutant
viral strains.1 The more effective regimen, highly active antiretrovi-
ral therapy (HAART), uses at least two anti-HIV drugs simulta-
neously. This therapy significantly reduces HIV viral load and has
led to a dramatic decrease in AIDS related mortality.2 The emer-
gence of HIV-1 strains resistant to at least one antiretroviral drug
highlights the need for further development of antiviral agents
with improved efficacy against these mutant strains.3
Reverse transcriptase (RT) inhibitors block the conversion of
single stranded viral RNA to double stranded DNA, a prerequisite
to integration into host DNA.4 Non-nucleoside reverse transcrip-
tase inhibitors (NNRTIs) interact with the RT enzyme at an alloste-
ric site to induce a change in the substrate binding site which
interferes with polymerase activity.5 A common consequence of
NNRTI use is the emergence of resistant mutant forms of HIV.6
Next generation NNRTIs are sought which possess both broad
spectrum antiviral activity against key mutant strains (especially
K103N and Y181C) and a high barrier to the selection of new mu-
tant strains. Additionally, facilitating patient compliance dictates
that next generation NNTRIs are efficacious with once daily oral
administration. The current leader in the pursuit of improved
NNRTI’s is Tibotec’s RilpivirineÒ (TMC278)7 which was recently ap-
proved by the FDA to be administered orally once daily.
In recent years, several groups have reported NNRTI’s which
feature a biaryl ether.8 Specifically, Tucker et. al. reported a series
of biaryl ethers (Fig. 1) which feature a pyrazolopyridine, as exem-
plified by MK-4965 (1), and have excellent broad spectrum antivi-
ral activity against key mutant strains of RT. We9 have also shown
that conformationally constrained inhibitors (Fig. 1) such as MK-
6186 (2) have comparable antiviral activity with good pharmacoki-
netics when dosed orally to preclinical species. Visual inspection of
the X-ray crystal structures of 1 (PDB entry 3DRP) and 3, a benzo-
triazole analog of 2 containing an aminomethyl group, complexed
to wt RT (Fig. 2) shows the importance of the positioning of the
pyrazolopyridine moiety with respect to the central phenyl ring
⇑
Corresponding author. Tel.: +1 215 652 4715; fax: +1 215 652 3971.
E-mail address: Robert_gomez@merck.com (R. Gomez).
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.10.027

R. Gomez et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7344–7350
7345
H2N
NH2
Cl
CN
Cl
CN
N
N
Cl
N
Cl
N
N
N
N
N
NH
NH
NH
O
O
O
O
N
N
N
Cl
Cl
Cl
1 (MK-4965)
2 (MK-6186)
3
Figure 1. Structures of lead NNRTI inhibitors.
W229
L234
W229
Y188
Y188
Y181
Y181
L234
L100
L100
V179
V179
V106
V106
F227
F227
K103
K103
P236
P236
Figure 2. The bound conformation of 3 (teal) (from crystal structure complex with
wt-RT) superimposed with the crystal structure of 1 (gray) complexed with wt-RT
(PDB entry 3DRP). Active site residues within 7 Å are displayed. Key active site
residues are highlighted and labeled in black. W239 and Y318 are hidden for clarity.
Figure 4. Docked conformation of 5 (salmon) superimposed with the bound
conformation of 1 (gray) complexed with wt RT. Active site residues within 7 Å are
displayed. Key active site residues are highlighted and labeled in black. W239 and
Y318 are hidden for clarity.
Cl
CN
F
Cl
CN
interactions with the K103 backbone in an analogous fashion to the
pyrazolopyridine of 1 and 3. We11 and others12 have shown that
one atom linkers between the pyrazolopyridine and the central
phenyl ring are also potent inhibitors of RT. In particular, the meth-
ylene linked 5 (Fig. 3) has sub-nanomolar enzyme inhibition
against both wt-RT and the two most clinically relevant mutants,
K103N and Y181C. Compound 5 was docked13 in the binding site
of 1 complexed with wt-RT. This docked conformation of 5 is
superimposed with the bioactive conformation of 1 (Fig. 4) and
shows exceptional alignment of the terminal rings. The linker to
the pyrazolopyridine of the docked conformation of 5 shows a
dihedral angle of 81° with respect to the central ring and indicates
the need for a nearly orthogonal disposition for a one atom linker
relative to the central core ring. This contrasts with the bound con-
formations of the 2 atom linked inhibitors, 1 and 3, which are more
planar with the linkers having measured dihedral angles of 150°
and 180° with respect to the central ring. 5 was unfortunately less
active in cell culture than 1, especially against the virus containing
the K103N mutant (211 nM), and had suboptimal pharmacokinet-
ics in rat. These shortcomings may be due to the nature of the cen-
tral phenyl ring and linker since the related compounds 1 and 2
have superior profiles. We sought to change the nature of the cen-
tral ring and linker of 5 in order to improve the antiviral activity in
cell culture and the pharmacokinetics in our preclinical species.
O
O
N
N
O
N
N
NH
NH
Br
Cl
5
4
RT inhibition EC50 (nM)
WT
K103N/Y181C 9
RT inhibition Ki (nM)
2
WT
0.24
0.48
0.51
K103N
Y181C
RT Antiviral Cell (CIC95 nM)
WT
K103N
22
211
Figure 3. Structures and RT inhibition of methylene linked NNRTI inhibitors.
as the resulting hydrogen bonds to the K103 backbone are essen-
tial for binding. The bound conformations of 1 and 3 complexed
with wt RT show dihedral angles of 150° and 180° with respect
to the linker between the pyrazolopyridine and the central phenyl
ring.
Sweeney et. al.10 have found that methylene linked inhibitors
such as 4 (Fig. 3) are potent inhibitors of RT. These compounds con-
tain a triazolinone moiety which presumably forms hydrogen bond

7346
R. Gomez et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7344–7350
Pyridones are a common platform used in drug development14
Cl
CN
Cl
CN
O
because their increased polarity is often more favorable than their
phenyl counterparts and the amide linkage leads to the possibility
of generating hydrogen bond interactions to increase affinity. Com-
putational analysis of N-alkyl pyridones15 shows a distinct confor-
mational preference in accordance with other ortho substituted
aromatics. A dihedral torsion driver was performed on N-ethyl
pyridone (blue curves) as shown in Figure 5. Minimization of the
steric interactions of the pyridone carbonyl with the ethyl group
leads to an energy minimum at a dihedral angle of 90°. This con-
trasts the synperiplanar disposition (dihedral angle of 0°) which
is predicted to be 6 kcal/mol higher in energy. The orthogonal
arrangement compliments the predicted bioactive conformation
of 5 from the docking study.12
A dihedral torsion driver was likewise performed on methoxy-
benzene15 to better understand the conformational preferences
of the oxymethylene linker of 1 as shown in Figure 5. Contrasting
the pyridone, the energy minima of methoxybenzene are at dihe-
dral angles of 0° and 180°. As anticipated, these planar dispositions
are the result of non-bonding lone pair electron conjugation with
the benzene ring. The observed dihedral angle of 150° for oxy-
methylene ether 1 complexed with wt RT is predicted to be only
0.5 kcal/mol from the energy minima at the dihedral angle 180°.
The central rings of 1 and 5 were replaced by a pyridone ring as
in Figure 6. We anticipated that the carbonyl oxygen atom of the
pyridone would be well tolerated as the benzotriazole nitrogen
atoms of 3 are accommodated in the binding site. The methylene
analog (n = 1) would be predicted from the dihedral torsion driver
of N-ethyl pyridone and docking studies of 5 to be a good match for
the bioactive conformation. The orthogonal placement is the low
energy conformation that is necessary for proper alignment of
the pyrazolpyridine with K103 backbone. The ethylene analog
(n = 2), however, was expected to be less compatible with the bio-
active conformation as the 150° dihedral angle optimal for proper
alignment of the pyrazolopyridine for two atom linkers would be
energetically unfavorable. The torsional strain for accessing the
bioactive conformation in this case is be predicted to be 3.0 kcal/
mol.
O
O
R
N
N
N
n
N
N
NH
NH
Pyridone analogs
Cl
5
Figure 6. Pyridone class of NNRTI inhibitors from 5.
group at the ortho position to direct the pyridine precursors toward
displacements at the required meta position. As such, 3-fluoro-4-
chloro pyridine (11) was deprotonated with lithium tetramethylpi-
peridide17 in hexanes and then quenched with bromine to give the
2-bromopyridine 12. This bromide was cyanated with zinc cyanide
and catalytic palladium(0) to provide the suitably activated 2-cya-
nopyridine 13. Heating 13 and 3-bromo-5-chlorophenol to 55 °C
with K2CO3 as base gave only the desired displacement product
14. The nitrile was then hydrolyzed to acid 15 using concentrated
HCl at 100 °C. Curtius rearrangement of the azide derived from car-
boxylic acid 15 using diphenyl phosphorylazide (dppa) with t-
butanol provided the Boc protected aminopyridine 16. This aryloxy
bromide was then cyanated using zinc cyanide in the presence of a
palladium(0) catalyst and the Boc group deprotected with TFA to
give the aminopyridine 17. The aminopyridine was converted to
the hydroxypyridine 18 via the hydrolysis of the corresponding
diazonium salt in sulfuric acid. This hydroxypyridine was selec-
tively N-alkylated with bromide 19 to provide, after deprotection,
8. The corresponding 4-methyl analog 6 was made in a similar
manner starting with the commercially available 2-bromo-3-flu-
oro-4-methylpyridine.
The synthetic approach used to prepare the ethylene linked
analog 7 is outlined in Scheme 2. The bromide 19 was treated with
NaCN in DMF to provide a mixture of products including the Boc
and des Boc desired product as well as by products derived from
alkylation of prematurely deprotected pyrazolopyridine. This
mixture was deprotected with TFA and the resulting mixture was
purified to provide 20. This nitrile was hydrolyzed with concd
HCl at 100 °C and the resulting carboxylic acid esterified in MeOH
to provide the ester 21. The pyrazolopyridine was protected with
THP by treatment with DHP and DDQ to provide 22. The methyl
ester was reduced with LAH in THF to provide the requisite
The chemistry was first examined in the context of the 4-
methyl pyridone analogs to afford 6 and 7. This synthetic approach
was then adapted to prepare the corresponding 4-chloro analog 8
as outlined in Scheme 1.16 The reactivity of 4-chloropyridines to-
ward SNAr reactions necessitated the introduction of an activating
ψ
O
ψ
N
ψ
ψ
O
ψ
ψ
Dihedral angle
ψ
Figure 5. Dihedral torsional energy drivers for N-ethylpyridone, ethylbenzene and methoxybenzene.

R. Gomez et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7344–7350
7347
Cl
Cl
Cl
Cl
Cl
O
a
c
b
F
F
F
H
N
Cl
Br
N
CN
N
11
N
12
Br
N
CN
Br
13
14
OH
f
Cl
Cl
Cl
Cl
Cl
Cl
O
d
e
O
O
N
CO2H
Br
NH(Boc)
N
NH2
N
16
Br
CN
15
17
Cl
CN
Cl
Cl
O
O
h
O
g
Cl
N
N
OH
CN
Br
N
Boc
18
8
N
N
N
HN
19
N
Scheme 1. Synthesis of 4-chloropyridone 8. Reagents and conditions: (a) BuLi, hexanes, À78 °C, then Br2, 39%; (b) Zn(CN)2, cat Pd(PPh3)4, DMF, 100 °C, 2 h, 40%; (c) K2CO3,
DMF, 55 °C, 20 min, 100%; (d) concd HCl, 100 °C, 2 h, 83%; (e) dppa, t-BuOH, pyridine, TEA, 65 °C, 1 h, 59%; (f) Zn(CN)2, cat Pd(PPh3)4, NMP, 90 °C, 1 h, then TFA, 71%; (g) NaNO2,
H2SO4, 87%; (h) K2CO3, DMF, 55 °C, 2 h, TFA, 92%.
HO
O
Br
NC
CN
a
O
b, c
d,e
N
N
N
N
N
N
N
N
H
N
N
N
H
O
N
Boc
19
20
21
22
CN
f
Cl
O
22
+
Cl
O
O
O
N
N
NH
N
7
NH
23
Scheme 2. Synthesis of ethylene linked 7. Reagents and conditions: (a) NaCN, DMF, rt, 2 h, then TFA, 20%; (b) concd HCl, 100 °C, 30 min, 66%; (c) HCl(g), MeOH, 86%; (d) DHP,
DDQ, ACN, 85 °C, 1 h, 90%; (e) LAH, THF, 0 °C, 76%; (f) DIAD, PPh3, THF then TFA, 18%.
hydroxyethyl pyrazolopyridine 23. Mitsunobu reaction of 23 with
the methylpyridone 24 followed by TFA deprotection provided 7.
The key synthetic steps to prepare 9 and 10 are outlined in
Scheme 3. The 4-chloro pyridone 18 was selectively chlorinated
using NCS in AcOH at 60 °C to give the bischloro pyridone 24. This
pyridone was alkylated with bromide 19 in DMF at 55 °C and then
deprotected with TFA to afford 9. The 6-aminopyrazolopyridine 10
was prepared from 18 via alkylation with chloride 25 and subse-
quent TFA deprotection at 75 °C.
Table 1 details the data for the pyridone analogs relative to the
oxymethylene linked 1. Compounds were evaluated for intrinsic
enzyme inhibitory potency18 versus wt RT as well as the K103N
and Y181C mutant variants. Compounds were also evaluated for
antiviral potency19 (CIC95) against wt and the key mutant viruses
in the presence of 10% fetal bovine serum (FBS) as well as with
50% normal human serum (NHS) to evaluate the effects of protein
binding.
The 4-methyl pyridone 6 was initially prepared due to its syn-
thetic tractability relative to the corresponding 4-chloro analog.
The 4-methyl pyridone 6 was 10-fold less potent in the enzyme
inhibition assay than MK-4965 (1) against wt RT and the clinically
relevant mutants, K103N and Y181C. Despite this loss in intrinsic
potency against RT, 6 had greater antiviral activity in cell culture
than 1 against wt, the K103N and Y181C mutant viruses as well
as the double mutant, K103N/Y181C. The improved ratio between
enzyme inhibition and antiviral activity in cell culture suggests
that the pyridone is less encumbered by non-specific binding. In
addition, the potency of 6 is shifted only 3.3-fold in the antiviral

7348
R. Gomez et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7344–7350
Cl
Cl
Cl
CN
Cl
Cl
CN
CN
O
O
a
Cl
Cl
b
O
Cl
O
N
N
OH
N
OH
N
HN
18
24
9
N
Cl
Cl
Cl
CN
Cl
CN
O
O
c, d
+
O
N
Cl
N
N
OH
PMB
N
25
N
H
N
PMB
18
N
HN
10
N
NH2
Scheme 3. Synthesis of compounds 9 and 10. Reagents and conditions: (a) NCS, AcOH, 60 °C, 2 h, 87%; (b) 19, K2CO3, DMF, 55 °C, 30 min, then TFA, 86%; (c) K2CO3, DMF, 55 °C,
2 h, then TFA, 75 °C, 2 h, 85% (2 steps).
Table 1
Enzyme inhibition and antiviral activity of pyridones 6–10 and compound 1
CN
CN
Z
Cl
O
Cl
O
Cl
X
Y
O
N
NH2
N
O
NH
N
N
Pyridone general structure
Antiviral activity in cell culture, CIC95 (nM)
1
NH
N
b
Compd
X
Y
Z
n
Inhibition of RT polymerase, Kia (nM)
WT
K103N
Y181C
WT
K103N
Y181C
K103N/Y181C
WT, 50%NHS
1
6
7
8
9
10
—
—
H
H
H
Cl
H
—
H
H
H
H
NH2
—
1
2
1
1
1
0.20
2.5
25
0.77
1.0
1.3
0.39
3.7
58
1.5
1.6
2.1
0.39
3.1
51
1.0
1.3
1.8
5.0
3.6
248
2.4
4.6
1.9
12
10
447
2.9
11
27
73
31
n.d.
7.0
31
28
12
611
7.7
15
Me
Me
Cl
Cl
Cl
n.d.
n.d.
7.8
15
7.1
63
30
11
a
Compounds were evaluated in a standard SPA assay. Values are the geometric mean of at least two determinations. Assay protocols are detailed in Ref. 18.
CIC95(cell culture inhibitory concentration) is defined as the concentration at which the Spread of virus is inhibited by >95% in MT-4 human T = lymphoid cells
b
maintained at RPMI 1640 medium containing either 10% FBS or 50% NHS. Details of the assay are provided in Ref. 19. Values are the geometric mean of at least 2
determinations. n.d. = not determined. No cytotoxicity was observed for any of the compounds up to the upper limit of the assay (8.3 M).
l
assay in the presence of normal human serum which is consistent
with it’s moderate plasma protein binding.
activity in cell culture. This loss of binding and antiviral activity
correlates with the increased torsional strain needed for the ethyl-
ene linked pyridone to adopt the bioactive conformation.
The chloro substituent was one of the most potent substituents
in the oxymethylene series21 and was incorporated into the
pyridone series. The resulting 4-chloro pyridone 8 was 3-fold more
potent an inhibitor of RT than 6 against both wt and the clinically
relevant mutants, K103N and Y181C. As a consequence, 8 had
significantly greater antiviral activity in cell culture than 6. Impor-
tantly, 8 had 10-fold more antiviral activity than 1 against the
K103N/Y181C double mutant virus.
The bis chloro pyridone 9 had slightly reduced intrinsic en-
zyme inhibition against wt and the K103N and Y181C mutant
viruses. The additional chloro group particularly affects cell activ-
ity leading to a 2-fold reduction in potency against the wt and
mutant viruses.
The crystal structure of 620 complexed with the K103N RT
(PDB code 3TAM) has been determined at a resolution of 2.5 ang-
stroms (Fig. 7). The terminal benzonitrile ring fills a large lipo-
philic pocket in the allosteric site formed by Y181, Y188, W229,
L234 and F227. The two faces of the pyridone ring interact with
V106 and L100, respectively, while the methyl containing edge
interacts with V179 and Y181. Importantly, the key backbone
interactions between K103 and the pyrazole nitrogens, which
are critical for binding, are maintained. Interestingly, the aryl
rings of the bioactive conformation of pyridone 6 are shifted
0.5–1.0 angstroms relative to the corresponding rings in the crys-
tal structure of biaryl ether 1.
The ethylene linked pyridone analog 7 was 10-fold less potent
than 6 in the enzyme inhibition assay and had 50-fold less antiviral

R. Gomez et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7344–7350
7349
mutant in which TMC-278 had an IC50 of >1
lM. This study dem-
onstrates that 8 possesses a broad spectrum of activity against a
variety of mutant viruses and appears to be an improved inhibitor
compared to some of the best next generation NNRTIs.
W229
Y188
Pyridone 8 was also evaluated for its antiviral potency of wt, the
K103N and Y181C mutants and the K103N/Y181C double mutant
viruses in 50% NHS and compared with 1, TMC-278 and UK-
453061. The use of 50% NHS is more relevant to the clinical envi-
ronment and reflects the expected influence of plasma protein
binding on the antiviral potency of these inhibitors. These antiviral
activities in cell culture using 50% NHS are illustrated in Figure 9.
For comparison, the potencies in 10% FBS are provided in parenthe-
ses. The shift in cell activity by changing from 10% FBS to 50% NHS
was considerably greater for TMC-278 (20–30-fold) relative to 1
(5–6-fold), UK-453061 (2-fold) and 8 (3–4-fold except) double mu-
tant) reflecting their relative affinities for plasma protein.24 Nota-
bly, the potency of 8 was superior to this collection of lead
inhibitors and had about 4-fold greater antiviral activity than the
other compounds when assayed in the presence of 50% NHS.
The pharmacokinetic parameters of the pyridone analogs was
determined in rats and are tabulated in Table 2. Sprague Dawley
rats received an intravenous (iv) dose as a solution in DMSO via
an indwelling canula implanted in the jugular vein. Animals re-
ceived an oral (p.o.) dose as a suspension in aqueous methylcellu-
lose. The plasma concentrations of the compounds were
determined by liquid chromatography mass spectral analysis and
the absolute oral bioavailability was determined by comparing
the mean area under the plasma concentration–time curve.
The pyridone class of inhibitors was characterized by achieving
only low exposure following p.o. administration. The lone excep-
tion was 8 which had exposures in rat comparable to 1. However,
the observed exposures showed high inter animal variation
Y181
L234
L100
V179
V106
F227
K103N
P236
Figure 7. The crystal structure of 6 (magenta) complexed with K103N mutant RT
(PDB entry 3TAM) superimposed with the bound conformation of 1 (gray) (from
PDB entry 3DRP). Key active site residues are highlighted and labeled in black.
Active site residues P99 and Y318 are hidden for clarity.
Analogously to work leading to 1,8 the 6-amino group was
incorporated onto the pyrazolopyridine of 8 to give compound
10. This compound was also slightly less potent in intrinsic enzyme
inhibition relative to 8 and also had significantly less antiviral
activity against the Y181C and K103N/Y181C mutant viruses.
The pyridone 8 was evaluated for its antiviral potency versus a
more extensive panel of clinically relevant mutant cell lines22,23
and compared with 1 (MK-4965), Tibotec’s TMC-278 and Pfizer’s
UK-453061. These activity profiles are illustrated in Figure 8 which
shows the various mutants versus their respective IC50 values for
each of the compounds in the study. The IC50 is defined as the con-
centration of compound in cell culture required to block 50% of vir-
al replication. Pyridone 8 in purple had a distinctly better profile
than both 1 (magenta) and UK-453061 (yellow). TMC-278 had a
comparable profile to 8 except for the K101P/K103N double
Antiviral activity in cell culture
with 50% NHS CIC95(nM)a
Cmpd
WT
K103N
Y181C
K103N/Y181C
8
7.7 (2.3) 12 (2.9)
20 (7.8)
104 (3.6)
68 (23)
69 (7.0)
311 (10)
100 (38)
589 (102)
TMC-278
36 (1.9) 46 (1.5)
UK-453061 40 (21)
70 (32)
54 (12)
1
26 (5)
175 (27)
Figure 9. Antiviral activity using 50% NHS in cell culture. aCIC95(Cell culture
inhibitory concentration) is defined as the concentration at which the Spread of
virus is inhibited by >95% in MT-4 human T = lymphoid cells maintained in RPMI
1640 medium containing 50% NMS. Details of the assay are provided in Reference #.
Values are the geometric mean of at least 3 determinations. No cytotoxicity was
observed for any of the compounds. Values in parenthesis are CIC95 using 10% FBS.
400
350
300
250
Table 2
8
IC50
200
150
100
50
Pharmacokinetics in rat of the pyridones 6–10 relative to 1 (MK-4965)
TMC-278
UK-453061
1
Compd Vd (L/kg) Clp (ml/min/kg) t1/2 (h) F %
AUCN (po) (lMÁh)
1b
6
2.0
3.7
0.6
2.0
27
0.8
9
11
31
12
27
12
3.5
11.7
0.75
4
19
1.4
52
8.5
—
57
0
2.3
0.31
—
7c
8
0
1.6a
0
9
10
6
0.15
Average of at least
2 Sprague Dawley rats dosed at 10 mpk PO (methocel
suspension) and 2 mpk IV (DMSO) unless otherwise noted. All values are within 25%
of the mean.
Virus (Mutation)
—
is not determined.
a
Variablity in exposures AUC = 24, 2.3, 1.5 and 38
l
M h to give a geometric mean
Figure 8. Antiviral potency of lead inhibitors versus panel of mutant cell lines.
Compounds were analyzed in a Monogram Bioscience Phenoscreen assay using 10%
FBS. The IC50 is defined as the concentration of compound in cell culture required to
block 50% of viral replication. Details provided in Ref. 23. WT R8 refers to a wild
type cell line.
of 16 15.5.
b
Average of 4 Sprague Dawley rats dosed at 10 mpk PO (methocel suspension)
and 3 mpk IV (DMSO).
c
Average of Sprague Dawley rats dosed at 1mpk IV (DMSO). No PO dosing in rats.

7350
R. Gomez et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7344–7350
M.; Suh, J. M.; Swallow, S.; Wu, J.; Hang, J. Q.; Zhou, A. S.; Klumpp, K. Bioorg.
Med. Chem. Lett. 2008, 18, 4348.
11. Tucker, T. J.; Tynebor. R. M. Unpublished results.
(>10-fold). This variability limits the potential of this compound for
development.
Next generation NNRTIs are being sought which possess both
broad spectrum antiviral activity against key mutant strains and
a high genetic barrier to the selection of new mutant strains. Pyri-
dones were evaluated as an acyclic conformational constraint to
replace the aryl ether core of MK-4965 (1) and the more rigid inda-
zole constraint of MK-6186 (2). The resulting pyridone class of
compounds were similarly potent inhibitors of HIV RT in vitro
but have superior antiviral activity in cell culture. This improved
antiviral activity was especially apparent when assayed in the
presence of 50% NHS. Further work aimed at increasing the bio-
availability of the pyridone inhibitors will be presented in due
course.
12. Sweeney, Z. K.; Harris, S. F.; Arora, N.; Javanbakht, H.; Li, Y.; Fretland, J.;
Davidson, J. P.; Billedeau, J. R.; Gleason, S. K.; Hirschfeld, D.; Kennedy_Smith, J.
J.; Mirzadegan, T.; Roetz, R.; Smith, M.; Sperry, S.; Suh, J. M.; Wu, J.; Tsing, S.;
Villasenor, A. G.; Paul, A.; Su Heilek, G.; Hang, J. Q.; Zhou, A. S.; Jernelius, J. A.;
Zhang, F.-J.; Klumpp, K. J. Med. Chem. 2008, 51, 7449.
13. The ligand 5 was docked within a 10 Å grid centroid of 1. The protein/ligand
complex was minimized using the low mode/mixed-torsional sampling
algorithm as implemented in Schrodinger, LLC v2009.1 with
dependent dielectric constant of 2 with the MMFFs force field.
a distance
14. Dragovitch, P. S.; Prins, T. J.; Zhou, R.; Johnson, T. O.; Hua, Y.; Luu, H. T.; Sakat, S.
K.; Brown, E. L.; Maldonado, F. C.; Tuntland, T.; Lee, C. A.; Fuhrman, S. A.;
Zalman, L. S.; Patrik, A. K.; Matthews, D. A.; Wu, E. Y.; Guo, M.; Borer, B. C.;
Nayyar, N. K.; Moran, T.; Chen, L.; Rejto, P. A.; Rose, P. W.; Guzman, M. C.;
Dovalsantos, E. C.; Lee, S.; McGee, K.; Mohajeri, M.; Liese, A.; Tao, J.; Kosa, M. B.;
Liu, B.; Batugo, M. R.; Gleeson, J. R.; Wu, Z. P.; Liu, J.; Meador, J. W.; Ferre, R. A. J.
Med. Chem. 2003, 46, 4572.
15. The dihedral torsion driver was performed using low mode/torsional sampling
Acknowledgments
every 10° as implemented in Schrodinger, LLC v2009.1 with
a distance
dependent dielectric constant of 1 with the OPLS.2005 force field. The dihedral
angle is formed by the relationship between the b atom of the linker and the
central ring.
We thank Dr. Steve Young for his guidance and mentorship of
this work. The authors also thank Sandor Varga and Mike Bogusky
for NMR support and Joan Murphy for exact mass determinations
in support of this project.
16. Full experimental details of the synthesis of compounds 6–10 are provided in
Tucker, T. J.; Tynebor, R.; Sisko, J. T.; Anthony, N. J.; Gomez, R.; Jolly, S. M. U.S.
Patent Appl. US 2007/3769P, 2007.
17. Marzi, E.; Bobbio, C.; Cottet, F.; Schlosser, M. Eur. J. Org. Chem. 2005, 10, 2116.
18. Saggar, S. A.; Sisko, J. T.; Tucker, T. J.; Tynebor, R. M.; Su, D.-S.; Anthony, N. J.
U.S. Patent Appl. US 2007/021442 A1, 2007.
References and notes
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20. The X-ray diffraction data of the K103N mutant RT and inhibitor 6 complex
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0.193 and an R-free of 0.249. The structure has been deposited in the Protein
Data Bank with a PDB code of 3TAM.
21. Tucker, T. J.; Saggar, S.; Sisko, J. T.; Tynebor, R. M.; Williams, T. M.; Felock, P. J.;
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versus a panel of clinically derived RT mutants in the presence of 10% fetal
bovine serum. The IC50 is defined as the concentration of compound in cell
culture required to block 50% of viral replication. Values are derived from the
average of two determinations. Assays were performed by Monogram
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bound.
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