3-Aryl-4-hydroxyquinolin-2(1H)-one derivatives as type I fatty acid synthase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2006.06.014

A series of 3-aryl-4-hydroxyquinolin-2(1H)-ones with fatty acid synthase inhibitory activity was prepared. Starting from a derivative with an IC₅₀ = 1.4 μM, SAR studies led to compounds with more than 70-fold increase in potency (IC₅₀

Full text of DOI:10.1016/j.bmcl.2006.06.014

Bioorganic & Medicinal Chemistry Letters 16 (2006) 4620–4623
3-Aryl-4-hydroxyquinolin-2(1H)-one derivatives as type I
fatty acid synthase inhibitors
Alexey Rivkin,a,* Yoona R. Kim,a Mark T. Goulet,a Nathan Bays,b
Armetta D. Hill,b Ilona Kariv,b Stefan Krauss,c Nicole Ginanni,c Peter R. Strack,c
Nancy E. Kohl,c Christine C. Chung,d Jeffrey P. Varnerin,d Paul N. Goudreau,d
Amy Chang,d Michael R. Totad and Benito Munoza
aDepartment of Chemistry, Merck Research Laboratories, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
bDepartment of Automated Lead Optimization, Merck Research Laboratories, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
cDepartment of Cancer Biology and Therapeutics, Merck Research Laboratories, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
dDepartment of Metabolic Disorders, Merck Research Laboratories, 126 Lincoln Avenue, Rahway, NJ 07065, USA
Received 10 April 2006; revised 1 June 2006; accepted 2 June 2006
Available online 19 June 2006
Abstract—A series of 3-aryl-4-hydroxyquinolin-2(1H)-ones with fatty acid synthase inhibitory activity was prepared. Starting from
a derivative with an IC50 = 1.4 lM, SAR studies led to compounds with more than 70-fold increase in potency (IC50 < 20 nM).
Ó 2006 Elsevier Ltd. All rights reserved.
Inhibition of human fatty acid synthase (FAS) has been
shown to induce apoptosis in a variety of cancer cells,
and consequently to be a potential therapeutic target
for the treatment of cancer.1 Expression and activity
of FAS is significantly higher in a variety of tumor cells
than in normal tissue, which creates the potential for a
large therapeutic index.2 To date, only a few inhibitors
of FAS have been reported and most of them are cova-
lent inhibitors.1d,e Of these the most commonly studied
are C75 (1) and cerulenin (2) which are covalent inhibi-
tors that have been shown to be efficacious in xenograft
studies at micromolar levels (Fig. 1).1d Recently, several
derivatives of the naturally occurring (5R)-thiolactomy-
cin (3) were shown to be reversible FAS inhibitors in the
micromolar range.3
a significant challenge in the development of small mol-
ecule inhibitors for human FAS.5
Herein, we report the synthesis and SAR studies of
3-aryl-4-hydroxyquinolin-2(1H)-one analogues as
a
novel class of FAS inhibitors. As part of our screen-
ing campaign, we identified 4, an interesting, albeit
weak, lead (IC50 value of 1.4 lM (human)). We rea-
soned that the enol functionality in both 3 and 4
may be mimicking the enol intermediate that occurs
during the enzymatic condensation reaction of
O
NH2
HO2C
O
O
O
O
FAS catalyzes the synthesis of long chain fatty acids
(primarily palmitate) from acetyl-CoA and malonyl-
CoA. The enzyme is a homodimer, each with seven cat-
alytic domains that in concert catalyze the reactions
leading to palmitate synthesis.4 The combination of
structural complexity and until recently the lack of
X-ray crystallography data of mammalian FAS creates
H3C(CH2)7
Cerulenin (2)
C75 (1)
O
S
HO
Keywords: Fatty acid synthase; Cancer; Apoptosis.
*
(5,R)-Thiolactomycin (3)
Corresponding author.Tel.: +1 617 992 2295; fax: +1 617 992 2403;
e-mail: alexey_rivkin@merck.com
Figure 1. Literature reported FAS inhibitors.
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.06.014

A. Rivkin et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4620–4623
4621
POBr3
or
acetyl-CoA and malonyl-CoA by the b-ketoacyl syn-
Br(Cl)
OH
thase domain of FAS. We subsequently initiated
SAR studies with 4 as our lead structure with the goal
of improving potency (see Fig. 2).
NC
Cl
NC
Cl
POCl3
DMF
N
H
O
110 o
C
N
H
O
11
12a,b
The general synthetic approaches to 3-aryl-4-hydroxy-
quinolin-2(1H)-one and variants are shown in Schemes
1–3. To explore the structure–activity relationship of
the quinolin-2(1H)-one core, we developed an efficient
and general solvent-free microwave cyclocondensation
reaction that is shown in Scheme 1.6 Irradiation of a
variety of electron-deficient or electron-rich anilines 5
with activated arylmalonate 6 led to the corresponding
products 7a–i in moderate to high yields. These com-
pounds were quickly purified by precipitation with
diethyl ether and filtration. Using this methodology we
were able to survey a variety of substituents on the quin-
olin-2(1H)-one core.
CuCN
R-NH2, DMSO,
DMF
150 oC, 16 h
NH
180 o
18h
C
R
CN
NC
Cl
NC
Cl
N
H
O
N
H
O
14
Scheme 3. Synthetic modifications of enol 11.
13a-c
To investigate the structure–activity relationship of the
phenyl moiety of 4, we utilized the traditional synthetic
approach to 3-aryl-4-hydroxyquinolin-2(1H)-one ana-
logues as shown in Scheme 2.7 Starting with methyl 2-
aminobenzoates 8, acylation with the appropriate acid
chloride in refluxing 1,2-dichloroethane gave the desired
corresponding phenylacetamides 9 in high yield. Intra-
molecular cyclization of the phenylacetamides 9 with
sodium bis(trimethylsilyl)amide in THF afforded the
desired 3-aryl-4-hydroxyquinolin-2(1H)-one analogues
10a–x in high yield. The hydroxy group could then be
converted to the corresponding bromide or chloride as
shown in Scheme 3. The bromide can then be displaced
with a variety of primary amines to provide the corre-
sponding enamines 13 or reacted with CuCN to give
the corresponding nitrile 14 (Scheme 3).
OH
O2N
N
H
4
O
Figure 2. Lead FAS inhibitor.
OH
μW, 250 o
15 min
C
R1
R2
R1
R2
NH2
N
H
O
R3
R3
The results of our initial SAR studies with rat fatty acid
synthase are shown in Table 1. The removal of the nitro
group led to complete loss of activity and migration of
the nitro group ortho to the nitrogen led to a 6-fold de-
crease in activity. Addition of a chloro or fluoro substi-
tuent ortho to the nitro group led to a remarkable 10-
fold improvement in potency (10b + 10c).
5
Cl
Cl
7a-i
O
O
O
O
Cl
Cl
Cl
Cl
6
Scheme 1. Microwave assisted synthesis of 3-aryl-4-hydroxyquinolin-
2(1H)-ones using activated aryl malonates.
We then turned our attention toward exploring the SAR
profile of the phenyl group (10e ! 10n) in hopes of find-
ing a permissive region. The removal of the phenyl
group led to complete loss of activity. Introduction of
fluoro or chloro substituents into the phenyl ring led
to a minor loss of activity. In contrast, replacement of
the phenyl group with 2-naphthyl or a variety of hetero-
cycles led to a substantial loss of activity. Finally, substi-
tution of the 4-position with a phenyl or benzyloxy
groups gave equally potent compounds.
O
O
O
R4
R1
R2
R1
R2
Cl
OMe
NH2
OMe
ClCH2CH2Cl
reflux, 16 h
NH
R3
R3
R4
O
8
9
NaHMDS
THF, rt, 1 h
Our SAR studies continued with human fatty acid syn-
thase (Table 2). Interestingly, in comparing the potency
of 7a and 10b against human and rat FAS, we observed
a moderate shift in in vitro potency. We were gratified to
find that the replacement of the nitro group with a cyano
group led to an improvement in activity, since the nitro
group represented a potential liability for toxicity. Intro-
duction of a chloride ortho to the cyano led to a 10-fold
increase in activity and the migration of the chloride to
OH
R1
R2
R4
N
O
H
R3
10a-x
Scheme 2. Traditional synthesis of 3-aryl-4-hydroxyquinolin-2(1H)-
ones.

4622
A. Rivkin et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4620–4623
Table 1. Rat fatty acid synthase inhibition data
of the binding site. Further substitution of the phenyl
group was well tolerated (10q–10x). The most favorable
substitution was a 2-methoxyphenyl group at the 3-posi-
tion (10v, IC50 = 19 nM), which represents a 74-fold
improvement in activity over the lead 4 (IC50 = 1.4 lM)
(see Table 3).
OH
R1
R2
R4
O
N
H
R3
R4
Compound R1
R2 R3
FAS
inhibition8
IC50 (nM)
In the final phase of our SAR studies we focused our ef-
forts on the substitution of the enol functionality (Table
4). Replacement of the 4-hydroxy group with a bromide,
chloride, amine, benzylamine, and nitrile group led to
complete loss of activity, which suggests that the 4-hy-
droxy group is essential for activity.
7a
7b
NO2
H
H
H
H
H
Ph
Ph
550
>15,000
3000
73
H
H
10a
10b
10c
10d
10e
10f
10g
10h
10i
10j
10k
10l
10m
10n
NO2 Ph
NO2 Cl
H
H
H
H
H
H
H
H
H
H
H
H
H
Ph
Ph
H
NO2
NO2
F
50
H
>15,000
443
173
In conclusion, we have discovered, through our SAR
studies, several 3-aryl-4-hydroxyquinolin-2(1H)-ones
that are potent human FAS inhibitors (IC50 = <100 nM)
against the human enzyme in vitro. These SAR studies
are highlighted by the successful replacement of the ni-
tro with nitrile moiety, which eliminated potential liabil-
ity for toxicity. In addition, this work led to the
identification of a permissive region in the 3-phenyl
group, which led to the discovery of some of our most
potent inhibitors.
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
NO2 Cl
2,4-Dichlorophenyl
2,4-Difluorophenyl
3,5-Dichlorophenyl
3,5-Difluorophenyl
2-Naphthyl
134
172
6000
700
2-Thienyl
1H-Tetrazol-5-yl
2600
3-Methylisoxazol-5-yl 3400
4-Biphenyl
4-(Benzyloxy)phenyl
122
86
Table 3. Human fatty acid synthase inhibition data
Table 2. Rat and Human fatty acid synthase inhibition data
OH
OH
3
NC
R1
R2
R
Cl
N
O
H
N
O
H
R3
Compound
R
H
FAS inhibition8
IC50 (nM)
Compound R1
R2 R3 Human FAS Rat FAS
inhibition8
IC50 (nM)
inhibition8
IC50 (nM)
7d
136
52
10o
10p
10q
10r
10s
10t
10u
10v
10w
10x
Phenyl
Methylsulfonyl
4-Fluorophenyl
3,4-Difluorophenyl
3,5-Difluorophenyl
2-Naphthyl
4164
54
7a
10b
7c
7d
7e
7f
NO2
H
Cl
H
Cl
H
H
H
H
H
H
H
H
H
Cl
H
H
H
H
1403
207
550
73
NO2
CN
CN
CN
CF3
68
1294
136
NA
NA
728
127
101
19
402
NA
1-Benzothien-3-yl
2-Methoxyphenyl
1-Methyl-1H-pyrazol-4-yl
>15,000
>15,000
>15,000
NA
>15,000
>15,000
>15,000
>15,000
7g
7h
7i
MeSO2
CF3SO2
CO2H
734
194
Pyridin-3-yl
NA, not available.
Table 4. Human fatty acid synthase inhibition data
the meta position led to a 4-fold loss in activity. Substi-
tution of the nitro group with other electron-withdraw-
ing groups such as trifluoromethyl, methyl sulfone,
trifluoromethyl sulfone, and carboxylic acid led to inac-
tive compounds.
R
NC
Cl
N
O
H
Compound
R
FAS inhibition8
IC50 (nM)
We then focused our efforts toward delineating the SAR
around the phenyl moiety of 7d. Interestingly, introduc-
tion of a methylsulfonyl group in the 3-position led to
40-fold loss in activity, whereas introduction of the
phenyl group in the same position led to a 3-fold
improvement in activity (10o vs 10p). This result sug-
gested that the 3-position occupies a hydrophobic region
12a
12b
13a
13b
13c
14
Br
Cl
>15,000
>15,000
>15,000
>15,000
>15,000
>15,000
NH2
p-MeO-Benzylamine
Benzylamine
CN

A. Rivkin et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4620–4623
4623
3. McFadden, J. M.; Medghalchi, S. M.; Thupari, J. N.; Pinn,
M. L.; Vadlamudi, A.; Miller, K. I.; Kuhajda, F. P.;
Townsend, C. A. J. Med. Chem. 2005, 48, 946.
4. Asturias, F. J.; Chadick, J. Z.; Cheung, I. K.; Stark, H.;
Witkowski, A.; Joshi, A. K.; Smith, S. Nat. Struct. Mol.
Biol. 2005, 12, 225.
References and notes
1. (a) Kuhajda, F. P.; Jenner, K.; Wood, F. D.; Hennigar, R.
A.; Jacobs, L. B.; Dick, J. D.; Pasternack, G. R. Proc. Natl.
Acad. Sci. U.S.A. 1994, 91, 6379; (b) Zhou, W.; Simpson, J.
P.; McFadden, J. M.; Townsend, C. A.; Medghalchi, S. M.;
Vadlamudi, A.; Pinn, M. L.; Ronnett, G. V.; Kuhajda, F.
P. Cancer Res. 2003, 63, 7330; (c) Knowles, L. M.; Axelrod,
F.; Browne, C. D.; Smith, J. W. A. J. Biol. Chem. 2004, 279,
30540; (d) Pizer, E. S.; Thupari, J.; Han, W. F.; Pinn, M. L.;
Chrest, F. J.; Frehywot, G. L.; Townsend, C. A.; Kuhajda,
F. P. Cancer Res. 2000, 60, 213; (e) Kuhajda, F. P.; Pizer,
E.; Li, J. N.; Mani, N. S.; Frehywot, G. L.; Townsend, C.
A. Proc. Natl. Acad. Sci. U.S.A. 2000, 97, 3450; (f) De
Schrijver, E.; Brusselmans, K.; Heyns, W.; Verhoeven, G.;
Swinnen, J. V. Cancer Res. 2003, 63, 3799.
2. (a) Pizer, E. S.; Lax, S. F.; Kuhajda, D. P.; Pasternack, G.
R.; Kurman, R. J. Cancer 1998, 83, 528; (b) Rashid, A.;
Pizer, E. S.; Moga, M.; Milgraum, L. Z.; Zahurak, M.;
Pasternack, G. R.; Kuhajda, F. P.; Hamilton, S. R. Am. J.
Pathol. 1997, 150, 201; (c) Milgraum, L. Z.; Witters, L. A.;
Pasternack, G. R.; Kuhajda, F. P. Clin. Cancer Res. 1997,
3, 2115.
5. Maier, T.; Jenni, S.; Ban, N. Science 2006, 311, 1258.
6. Rivkin, A.; Adams, B. Tetrahedron Lett. 2006, 47, 2395.
7. (a) DeVita, R. J.; Hollings, D. D.; Goulet, M. T.; Wyvratt,
M. J.; Fisher, M. H.; Lo, J. L.; Yang, Y. T.; Cheng, K.;
Smith, R. G. Bioorg. Med. Chem. Lett. 1999, 9, 2615; (b)
DeVita, R. J.; Goulet, M. T.; Wyvratt, M. J.; Fisher, M. H.;
Lo, J. L.; Yang, Y. T.; Cheng, K.; Smith, R. G. Bioorg.
Med. Chem. Lett. 1999, 9, 2621.
8. In vitro enzymatic activity of FAS was determined by
monitoring the production of free coenzyme A (CoA-SH)
from malonyl- and acetyl-CoA during the FAS enzymatic
reaction. CoA-SH levels were determined using the thiol-
reactive dye 7-diethylamino-3-(4-maleimidophenyl)-4-
methylcoumarin (CPM). Unreacted CPM is nonfluo-
rescent, whereas the CPM–CoA adduct shows a peak
fluorescence emission at 475 nM after excitation at
380 nM.