Substituted coumarins as potent 5-lipoxygenase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2006.01.085

Leukotriene biosynthesis inhibitors have potential as therapeutic agents for asthma and inflammatory diseases. A novel series of substituted coumarin derivatives has been synthesized and the structure-activity relationship was evaluated with respect to th

Full text of DOI:10.1016/j.bmcl.2006.01.085

Bioorganic & Medicinal Chemistry Letters 16 (2006) 2528–2531
Substituted coumarins as potent 5-lipoxygenase inhibitors
Erich L. Grimm,* Christine Brideau, Nathalie Chauret, Chi-Chung Chan, Daniel Delorme,
Yves Ducharme, Diane Ethier, Jean-Pierre Falgueyret, Richard W. Friesen, Jocelyne Guay,
Pierre Hamel, Denis Riendeau, Chantal Soucy-Breau, Philip Tagari and Yves Girard
Merck Frosst Centre for Therapeutic Research, 16711 Trans Canada Hwy, Kirkland, Que., Canada H9H 3L1
Received 2 December 2005; revised 19 January 2006; accepted 19 January 2006
Available online 7 February 2006
Abstract—Leukotriene biosynthesis inhibitors have potential as therapeutic agents for asthma and inflammatory diseases. A novel
series of substituted coumarin derivatives has been synthesized and the structure–activity relationship was evaluated with respect to
their ability to inhibit the formation of leukotrienes via the human 5-lipoxygenase enzyme.
Ó 2006 Elsevier Ltd. All rights reserved.
5-Lipoxygenase (5-LO) is a key enzyme in the biosynthe-
sis of leukotrienes and catalyzes the initial steps in con-
version of arachidonic acid to leukotrienes.1,2 Inhibition
of this enzyme may decrease leukotriene-mediated
inflammatory responses and control disease states such
as asthma.3,4 Recent studies have implicated 5-LO activ-
ity5–7 in a number of other diseases, including COPD,
cancer, osteoporosis, and atherosclerosis, and several re-
views have appeared recently highlighting the therapeu-
tic potential of 5-LO inhibition.8–14
octanyl moiety and precluded further development of
these inhibitors.17
We therefore set out to identify replacements for the
dioxabicyclooctanyl subunit as well as for the furyl sub-
stituent18 in an effort to reduce the potential for toxicity.
This work has culminated in the discovery of inhibitor 1
in which the 2-cyanoquinoline portion of L-746,530 was
replaced by a fluorophenyl-substituted coumarin and
the dioxabicyclooctanyl moiety by a hexafluorocarbinol
substituent.
Many of the currently described inhibitors of 5-LO con-
tain functional groups such as phenol, hydroxamate or
N-hydroxyurea and act by a redox mechanism or by
chelation of the active site iron. The multiple toxicities
and difficulties encountered in developing redox inhibi-
tors of 5-LO have led many research groups to strive
to find competitive non-redox inhibitors of this enzyme.
From previous structure–activity relationship (SAR)
studies we learned that the 2-cyanoquinoline moiety in
L-746,530 could be replaced by a 4-substituted couma-
rin.19 This modification was a welcome observation
F
We have previously introduced new classes of non-redox
5-LO inhibitors such as the pyridyl-substituted 2-cyano-
naphthalene L-739,01015 and the phenyl-substituted 2-
cyanoquinoline L-746,53016 (Fig. 1). While both inhibi-
tors have excellent potency and pharmacokinetics in ani-
mal models, microsomal incubation studies revealed the
formation of reactive intermediates at the dioxabicyclo-
O
O
CN
N
O
N
CN
OH
O
O
OH
O
O
F
L-746,530
L-739,010
O
F3C
F3C
S
O
O
OH
Keywords: 5-Lipoxygenase; Leukotrienene inhibitor; Coumarin;
Structure–activity relationship.
1
F
*
Corresponding author. Tel.: +514 428 3025; e-mail: erich_grimm@
merck.com
Figure 1.
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.01.085

E. L. Grimm et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2528–2531
2529
since, in theory, this would enable the preparation of
coumarin prodrugs that could be delivered orally as
water-soluble hydroxy acids that would cyclize in vivo
to the active coumarin derivative (vide infra). All com-
pounds prepared were evaluated for their potency to
inhibit the oxidation of arachidonic acid by recombinant
human 5-lipoxygenase (H5-LO),20 the production of
LTB4 in calcium ionophore-stimulated human periphe-
ral blood polymorphonuclear leukocytes (HPMN),21
and the production of LTB4 in calcium ionophore-stim-
ulated human whole blood (HWB).21 Although 10-fold
less potent on H5-LO, no significant loss of inhibitory
potency in HWB was observed by replacing the 2-cyano-
quinoline in reference compound L-746,530 with cou-
marin 2 (IC50 = 48 nM, Table 1, entry 2). The phenyl
derivative 3 and the 4-fluorophenyl derivative 4, howev-
er, were significantly less potent in the human whole
blood assay with IC50s of 440 and 1700 nM, respectively
(Table 1, entries 3 and 4).
In particular, the metabolically unstable dioxabicyclooc-
tanyl subunit of 2 could be replaced by a variety of sub-
stituents (Table 2, entries 1–4). For example, the 2-
thiazolyl ethyl derivative 522 (racemic) (entry 1) and
the 2-pyridinyl ethyl derivative 6 (racemic) (entry 2)
are tolerated, while a small gain in potency in HWB
(IC50 = 89 nM) can be achieved by replacing the hetero-
cycle with an additional ethyl substituent to give 7 (entry
3). Although 7 exhibited excellent in vitro potency, it
suffered from poor pharmacokinetics presumably due
to metabolism at the geminal diethyl substituent. This
problem could be alleviated by replacing the ethyl sub-
stituents with a geminal trifluoromethyl group to give
8 (entry 4). This bis(trifluoromethyl)carbinol moiety
was found to be an excellent surrogate for the metabol-
ically unstable dioxabicyclooctanyl alcohol and, there-
fore, was used exclusively in optimization studies.
In our earlier work, we demonstrated that the nature of
the linkage (oxymethylene link vs thio link) between two
aromatic moieties can have a profound effect on the
potency of 5-LO inhibitors.16 Therefore, it was pertinent
to examine this structural modification in this series (Ta-
ble 3). With the exception of the p-chloro-substituted
inhibitor 9 (Table 3, entry 2), the furyl derivative 10,
One of the most interesting avenues of SAR that was
pursued, and that led to a wide range of inhibitors with
acceptable potency, was derived from the preparation of
different tertiary alcohols (Table 2).
Table 3. SAR of hexafluorocarbinol-substituted coumarins
Table 1. SAR of dioxabicyclooctanyl-substituted coumarins
F
F
F3C
O
O
R
O
O
F3C
S
O
R
O
OH
O
OH
a
Entry Compound
R
IC50 (nM)
H5-LOb HPMNb HWBb
27 10 2.3 1.1 36 18
3-Furyl 200 70 3.0 0.9
Ph 300 75 3.7 0.6
4-F–Ph n.d.c
a
IC50 (nM)
Entry Compound
R
H5-LOb
HPMNb HWBb
1
2
3
4
L-746,530
1
2
3
4
1
9
4-F–Ph
27 16 0.5 0.3
70 24
4-Cl–Ph 180 42 4.3 3.7 400 45
2
3
4
48 17
440 150
11.6 0.1 1700d
10
11
3-Furyl
Ph
9
2.0 0.8
0
26 19 0.5 0.2
52 21
88 21
a Mean SD.
b n P 2.
a Mean SD.
b n P 2.
c Not determined.
d n = 1.
Table 2. SAR of carbinol-substituted coumarins
F
R1
R2
O
O
O
OH
R3
R1
R2
R3
a
IC50 (nM)
Entry
Compound
H5-LOb
HPMNb
HWBb
1
2
3
4
5
6
7
8
Et
Et
Thiac
Pyrc
Et
4-F–Ph
3-Furyl
3-Furyl
4-F–Ph
175 20
175 25
15 5.0
55 4.0
2.7 3.2
3.7 1.7
0.9 0.6
1.9 0.5
400 10
360 20
89 5.0
150 75
Et
CF3
CF3
a Mean SD.
b n P 2.
c Thia = 2-thiazolyl, Pyr = 2-pyridinyl.

2530
E. L. Grimm et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2528–2531
HO
OH
the phenyl derivative 11, and the p-fluorophenyl deriva-
tive 1 (entries 3, 4, and 1) showed excellent inhibitory
potency in all in vitro assays.
HO
O
O
a
F
+
MeO
Bioavailability studies with 1 were performed by admin-
istering the water-soluble sodium salt of the correspond-
ing hydroxy acid 1223 (Scheme 1).
O
O
F
13
Br
O
O
b
This compound, when dosed orally in rats at 20 mg/kg
in 0.5% methocel, was well absorbed and readily con-
verted to active 1.24 Thus, upon dosing 12, excellent
bioavailability (F = 100%) (0–24 h) and a maximum
concentration (Cmax) of 16–21 lM (1 and 4 h after dos-
ing) were observed for 1. Interestingly, oral administra-
tion of 1 as a suspension (20 mg/kg in 0.5% methocel)
did not result in any absorption at all presumably due
to low solubility. Therefore, 12 was used in all bio-
availability studies. In squirrel monkeys, when 12 was
dosed orally at 10 mg/kg in 0.5% methocel, bioavail-
ability of 30% for 1 can be achieved with a Cmax of
1.2 lM at 2 h. The inhibitory effect of 1 on the biosyn-
thesis of LTB4 in vivo was evaluated using the rat pleu-
ral cavity model following carrageenan-induced
inflammation.25 In this model, 1 showed activity with
an ED50 of 1.6 mg/kg 2 h after oral dosing of 12.
The potency of 1 on: (1) the inhibition of urinary
LTE4 (an index of the systemic biosynthesis of peptido-
leukotrienes) and (2) the ex vivo generation of LTB4 in
whole blood stimulated with calcium ionophore
A23187 was measured in an anesthetized dog model.26
1 inhibited base-line urinary LTE4 excretion, measured
5–7 h after the commencement of the infusion of 1 in
80% PEG200/water with an ED50 of 2.5 lg/kg/min.
Compound 1 also inhibited the production of LTB4
in dog whole blood ex vivo with an ED50 of 2.5 lg/
kg/min. Microsomal metabolism studies with radiola-
beled 127 showed that there was no formation of radio-
active metabolites covalently bound to protein, a
problem we had encountered with our previous inhibi-
tors L-737,010 and L-746,530.17
F
14
15
F
F
c
Br
F
Br
SC(Me)3
F
F
d
e
F3C
F3C
F3C
F3C
SC(Me)3
SH
OH
OH
16
17
F
F3C
F3C
f
S
O
F
O
OH
1
Scheme 2. Reagents and conditions: (a) HCl, MeOH, 0 °C to rt, 15 h
(79%); (b) Br2, PPh3, 280 °C, 1 h (62%); (c) Me3CSH, NaH, DMF,
À10 °C to 0 °C, 24 h (78%); (d) Mg, CF3COCF3, THF, 0 °C, 1.25 h
(72%); (e) 1—Hg(OAc)2, PhOMe, TFA, DMF, 60 °C, 3.5 h; 2—
Na2SÆ7H2O, rt, 0.25 h (56%); (f) K2CO3, NMP, 100 °C, 1 h (66%).
methyl-2-propanethiol.30 The coupling product 15, ob-
tained in 78% yield, was then engaged in a Grignard
reaction with liquified hexafluoroacetone to yield 16 in
72% yield. Deprotection of 16 with mercury(II) acetate
and sodium sulfide31 gave thiol 17 in 56% isolated yield.
Final coupling of 17 and 14 with potassium carbonate in
1-methyl-2-pyrrolidinone (NMP) furnished 1 in 66%
yield as an off-white solid.
The synthesis of 1 was accomplished (Scheme 2) in a
convergent manner by the preparation and coupling of
two advanced intermediates, the aromatic thiol 17 and
the bromocoumarin 14. Preparation of bromide 14 com-
menced with a von Pechmann condensation28 of resor-
cinol and methyl 4-fluorobenzoylacetate under acidic
conditions to provide phenol 13 in 79% yield. Treatment
of 13 with bromine (neat) and triphenylphosphine at
280 °C furnished 14 in 62% isolated yield.29 Thiol 17
was prepared in a four-step sequence starting with com-
mercially available 1-bromo-3,5-difluorobenzene and 2-
The results of the present study demonstrate that the
structures of L-739,010 and L-746,530 can be modified
to yield 5-LO inhibitors with comparable in vitro poten-
cy. With the bis(trifluoromethyl)carbinol moiety we
have identified an alternative motif for the dioxabicyclo-
octanyl ring system. In addition, we have replaced the
furyl-substituted 2-cyanoquinoline with a 4-fluorophe-
nyl coumarin and the optimized 5-LO inhibitor 1 has
excellent pharmacokinetics in rats when dosed as the
prodrug 12.32
F
F
F3
F3C
C
ONa
CO2Na
F3C
F3
S
O
F
O
S
C
OH
OH
References and notes
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F
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Scheme 1.

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