6-Alkyl, alkoxy, or alkylthio-substituted 3-(4-methanesulfonylphenyl)-4-phenylpyran-2-ones: A novel class of diarylheterocyclic selective cyclooxygenase-2 inhibitors

Article abstract of DOI:10.1016/S0960-894X(03)00391-3

A novel class of 3-(4-methanesulfonylphenyl)-4-phenylpyran-2-ones possessing a central six-membered lactone (pyran-2-one) ring system, in conjunction with C-6 alkyl (Me, Et or i-Pr), alkoxy (OMe, OEt or O-i-Pr), and alkylthio (SMe, SEt or S-i-Pr) substitu

Full text of DOI:10.1016/S0960-894X(03)00391-3

Bioorganic & Medicinal Chemistry Letters 13 (2003) 2205–S2209
6-Alkyl, Alkoxy, or Alkylthio-Substituted
3-(4-Methanesulfonylphenyl)-4-phenylpyran-2-ones:
A Novel Class of Diarylheterocyclic Selective
Cyclooxygenase-2 Inhibitors
P. N. Praveen Rao, Mohsen Amini, Huiying Li,
Amgad G. Habeeb and Edward E. Knaus*
Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta,
Edmonton, Alberta, Canada T6G 2N8
Received 6 January 2003; accepted 21 March 2003
Abstract—A novel class of 3-(4-methanesulfonylphenyl)-4-phenylpyran-2-ones possessing a central six-membered lactone (pyran-2-
one) ring system, in conjunction with C-6 alkyl (Me, Et or i-Pr), alkoxy (OMe, OEt or O-i-Pr), and alkylthio (SMe, SEt or S-i-Pr)
substituents, were designed for evaluation as selective COX-2 inhibitors.
# 2003 Elsevier Science Ltd. All rights reserved.
Selective cyclooxygenase-2 (COX-2) inhibitors currently
provide effective treatment of inflammatory disease
states such as rheumatoid arthritis and osteoarthritis.1
Recent studies have shown that selective COX-2 inhibi-
tors can also induce apoptosis in colon, stomach, pros-
tate and breast cancer cell lines.2 Selective COX-2
inhibitors offer potential for the prophylactic prevention
of inflammatory neurodegenerative disorders such as
Alzheimer’s disease.3
belongs to a diarylheterocylic class that possesses a
central five-membered lactone, [2(5H)furanone], ring
system.7 We describe herein the design, synthesis and
biological evaluation of a novel class of diarylhetero-
cyclic, 6-alkyl, alkoxy or alkylthio-substituted-3-(4-
methanesulfonylphenyl)-4-phenylpyran-2-ones,
that
possesses a central six-membered lactone (pyran-2-one)
ring.
Diarylheterocycles constitute a major class of selective
COX-2 inhibitors. In this regard, celecoxib (1) possesses
a central five-membered pyrazole ring, whereas eto-
ricoxib (2) has a central six-membered pyridine ring.4
Extensive structural–activity relationship (SAR) studies
for the diarylheterocycle class have shown that a
SO2NH2 or SO2Me and F substituents at the para-
position of one of the aryl rings often provides optimum
COX-2 selectivity and potency.5 Thus, the selective
COX-2 inhibitor SC 57666 (3) has a sulfonylmethyl
group at the para-position of one phenyl ring along with
a fluorine atom at the para-position on the other phenyl
ring.6 The highly selective COX-2 inhibitor rofecoxib (4)
*Corresponding author. Tel.: +1-780-492-5993; fax: +1-780-492-
1217; e-mail: eknaus@pharmacy.ualberta.ca
0960-894X/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0960-894X(03)00391-3

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P. N. P. Rao et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2205–S2209
The synthetic reactions used for the synthesis of 6-alkyl,
alkoxy or alkylthio-substituted-3-(4-methanesulfonyl-
phenyl)-4-phenylpyran-2-ones (10a–c, 11a–c, and 12a–c)
are outlined in Schemes 1–3. The 2,3-diphenylcyclopro-
penone (6) with a thiomethyl substituent at the para-
position of one of the phenyl rings was prepared in
moderate yields (22–33%) using a one-pot reaction
starting with tetrachlorocyclopropene (5). The sequen-
tial arylation of 5 with benzene and methylthiobenzene,
followed by hydration with ice-water yielded 2-(4-
methylthiophenyl)-3-phenylcycloprop-2-en-1-one (6) as
the major product along with the 2-(2-methylthiophe-
nyl)-3-phenylcycloprop-2-en-1-one regioisomer as
a
Scheme 2. Reagents and conditions: (a) pyridine, THF, 25 ꢀC, 6 h.
minor roduct (ratio 4:1) which could not be purified by
column chromatography. Subsequent oxidation of 6
using aqueous Oxone1 solution afforded the methane
sulfonyl product 7 as shown in Scheme 1.8 The N-alkyl,
alkoxy or alkylthiocarbonylmethyl pyridinium chlorides
or bromides (9a–i, 33–68%) were prepared by the reac-
tion of the respective alkyl, alkoxy or alkylthio-sub-
stituted chloro or bromoacetate derivatives (8a, 8b, or
8c) with pyridine as illustrated in Scheme 2.
The target 6-alkyl, alkoxy or alkylthio-substituted-3-(4-
methanesulfonylphenyl)-4-phenylpyran-2-ones (10a–c,
11a–c, and 12a–c) were prepared in low to moderate
yields (9–44%) by the condensation of 7 with the
respective pyridinium chlorides or bromides (9a–c, 9d–f,
or 9g–i) in the presence of the base Et3N to produce an
intermediate ylide product that undergoes a ring
expansion reaction to afford the title products (10a–c,
11a–c, or 12a–c) as illustrated in Scheme 3.9 The struc-
tures of 10–12 were confirmed by microanalyses data
Scheme 3. Reagents and conditions: (a) benzene, triethylamine, 25 ꢀC,
16–18 h.
1
and H NMR NOE studies which showed NOE inter-
actions between H-5 and the 6-alkyl (R), alkoxy (O-R)
or alkylthio (S-R) moiety, and between H-5 and the C-4
ortho-phenyl hydrogens, which establishes the regio-
chemistry of the C-4 and C-5 phenyl rings.
IC50=281.5 mM; COX-2 IC50=1.3 mM; COX-2 Selec-
tivity Index=216.5). Introduction of a thioethyl (EtS-)
substituent at C-6 of the central pyran-2-one ring led to
a dramatic increase in COX-2 selectivity and potency,
with 12b showing a weak COX-1 inhibition (COX-1
IC50=386.2 mM) and potent inhibition of COX-2
(COX-2 IC50=0.0032 mM) for a very high COX-2 S.I.
>120,000 relative to the reference drug rofecoxib
(COX-2 IC50=0.4279; S.I. >1168).
The effect of the C-6 alkyl (10a–c), alkoxy (11a–c), and
alkylthio (12a–c) substituents on the central six-mem-
bered lactone (pyran-2-one) ring on COX-2 selectivity
and potency was determined by in vitro COX-1/COX-2
inhibition studies. The structure–activity relationships
acquired showed that this lactone class of compounds
are moderate to potent selective COX-2 inhibitors (see
data in Table 1).
The critical difference between the binding sites for
COX-1 and COX-2 is at position 523 where COX-2 has
the amino acid residue Val in place of the bulkier Ile in
COX-1. This difference produces a secondary pocket
extending off the primary binding site in COX-2 that is
absent in COX-1. Consequently, the combined volume
of the primary binding site and the secondary pocket in
COX-2 is about 25% larger (394 A3) than the volume of
The
6-alkyl-3-(4-methanesulfonylphenyl)-4-phenyl
pyran-2-ones (10a–c), show weak to moderate COX-1
inhibition (8.0–614.8 mM range) with good COX-2 inhib-
ition in the 0.5–1.5 mM range. In the alkoxy series (11a–
c), good COX-2 inhibitory activity and selectivity was
shown by the 6-ethoxy derivative 11b (COX-1
Scheme 1. Reagents and conditions: (a) dry AlCl3, 1,2-dichloroethane, benzene, 25 ꢀC, 24 h; (b) thioanisole, 25 ꢀC, 24 h; (c) H2O, 25 ꢀC, 10 min; (d)
aqueous Oxone1, THF–MeOH (1:1), 25 ꢀC, 4–5 h.

P. N. P. Rao et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2205–S2209
2207
Table 1. In vitro inhibition of COX-1 and COX-2 by 6-alkyl, 6-
alkoxy or 6-alkylthio-3-(4-methanesulfonylphenyl)-4-phenylpyran-2-
ones (10a–c, 11a–c, and 12a–c)
placement of substituents with varying electronic and
steric properties.11
It is well established for the diarylheterocyclic class of
COX-2 inhibitors, that a para-methylsulfone or sulfo-
namide substituent on one of the phenyl rings is a
requirement for good COX-2 potency and selectivity.5
Accordingly, the 6-alkyl, 6-alkoxy, and 6-alkylthio-3-(4-
methanesulfonylphenyl)-4-phenylpyran-2-one group of
compounds were designed to have a –SO2Me sub-
stituent at the para-position of one of the phenyl rings.
Compounds 10a–c, 11a–c, and 12a–c have volumes in
the range of 293–345 A3, relative to the selective COX-2
inhibitor rofecoxib (267.2 A3) as shown in Table 1. In
general, for this series of compounds, COX-2 selectivity
and potency was dependant upon steric and electronic
properties of the C-6 substituent on the central pyran-2-
one ring which positions the sulfonylmethyl moiety in
the vicinity of the secondary pocket of COX-2.
Compd
COX-1
inhibition
IC50, mMa
COX-2
inhibition
IC50, mMa
COX-2
S.I.b
Volume
(A3)c
10a
10b
10c
11a
11b
11c
12a
12b
614.8
8.0
341.5
14.7
281.5
4.0
>100
386.2
>100
>500
22.9
0.68
1.5
0.50
28.3
1.3
2.0
2.8
904.0
5.3
683.0
<0.52
216.5
2.0
293.56
310.18
326.69
301.69
318.75
336.04
311.54
328.56
345.65
267.20
298.56
35.7
120,687.5
0.0032
>100
0.4279
0.0567
12c
Rofecoxib
Celecoxib
>1,168
404
aValues are means of two determinations acquired using an ovine
COX-1/COX-2 assay kit (Catalog No. 560101, Cayman Chemicals
Inc., Ann Arbor, MI) and the deviation from the mean is <10% of
the mean value.
bIn vitro COX-2 selectivity index (IC50 COX-1/IC50 COX-2).
cThe volume of the molecule, after minimization using the MM3 for-
cefield, was calculated using the Alchemy 2000 program.
The orientation of the highly potent and selective COX-
2 inhibitor, 6-ethylthio-3-(4-methanesulfonylphenyl)-4-
phenylpyran-2-one (12b), in the COX-2 active site was
examined by a docking experiment (Fig. 1).12 This study
showed that 12b binds in the center of the primary
binding site of COX-2 with the SO2Me moiety interact-
ing with the secondary pocket amino acid residues
Phe518, Gln192, Arg513, Leu352, Ser353 and Val523. One of
the COX-1 binding site (316 A3).10 This difference in
volume can be exploited to manipulate COX-2 selectiv-
ity of diarylheterocyclic classes of COX-2 inhibitors, by
varying the volume of the drug and the appropriate
Figure 1. Docking of 12b (ball and stick) in the active site of murine COX-2 (line and stick) (Eintermolecular=ꢁ90.81 kcal/mol). Hydrogen atoms of
the amino acid residues are removed to increase clarity.

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P. N. P. Rao et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2205–S2209
the O-atoms of the SO2Me substituent forms a hydro-
gen bond with the amide hydrogen of Phe518 (1.92 A).
The ring O-atom of the central lactone (pyran-2-one) is
oriented in the direction of the polar amino acid Arg120
at the mouth of the channel, where this O-atom is about
4.24 A away from the NH2 (guanidino) group. The
C¼O of the central pyran-2-one is hydrogen bonding
with the OH of Tyr355 (1.70 A). These interactions may
disrupt the salt bridge between His90, Arg120, Tyr355 and
Glu524 at the mouth of the COX-2 active site. The
unsubstituted phenyl ring lies in a hydrophobic cavity
lined by Tyr385, Trp387, Tyr348 and Ser530. Interestingly,
the C-6 EtS-substituent is located in a hydrophobic
region formed by Val344, Ile345, Val349, Ser530 and
Leu531, with the S-atom forming a weak hydrogen bond
with the OH of Ser530 (4.41 A). This shows the impor-
tance of the C-6 substituent in orienting the molecule
such that the methylsulfone moiety inserts into the sec-
ondary pocket of COX-2. A similar docking study for
the less potent, and less selective, COX-2 inhibitory C-6
OEt analogue (11b) showed that the SO2Me moiety is
inserted less deeply into the secondary pocket than the
C-6 SEt of 12b, the lactone ring oxygen atom in 11b is
closer to the NH2 of Arg120 (3.26 A) relative to 4.24 A in
12b, that the C-6 OEt oxygen atom is not within
hydrogen bonding distance of the OH of Ser530 (6.63 A),
and the intermolecular energy for the ligand-enzyme
complex for 11b is higher (ꢁ87.60 kcal/mol). These
observations together with the larger volume (328.5 A3),
provides a good explanation for the potent and selective
inhibitory activity of 12b.
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The results of this investigation show (i) a C-6 SEt sub-
stituent (12b)13 in this 3-(4-methanesulfonylphenyl)-4-
phenylpyran-2-one class of diarylheterocycles provides
potent and selective inhibition of the COX-2 isozyme,
(ii) molecular modeling studies indicate the SO2Me
moiety inserts deep into the COX-2 secondary pocket
and the C-6 SEt sulfur atom forms a weak hydrogen
bond with the OH atom of Ser530 and (iii) these C-6
alkyl, alkoxy and alkylthio compounds 10–12 could
serve as useful probes to study the function and cataly-
tic activity of the COX-2 isozyme.
12. Docking studies were performed using Insight II software
Version 2000.1 (Accelrys Inc.) running on a Silicon Graphics
Octane 2 R14000A workstation. The coordinates of the X-ray
crystal structure of the selective COX-2 inhibitor SC-558
bound to the murine COX-2 enzyme was obtained from the
RCSB Protein Data Bank (1cx2) and hydrogens were added.
The ligand molecules were constructed using the Builder
module and were energy minimized for 1000 iterations reach-
ing a convergence of 0.01 kcal/mol A. The energy minimized
ligands were superimposed on SC-558 in the PDB file 1cx2
after which SC-558 was deleted. The resulting structure
(ligand–enzyme assembly) was minimized using the Discover
module for 5000 iterations or until an RMSD of 0.05 A was
reached using consistent valence force field (CVFF). Further
optimization of the ligand–enzyme complex was obtained
using the Affinity command in the Docking module of Insight
II by defining subsets of the enzyme such that residues within
10 A of the ligand were allowed to relax, while the rest of the
enzyme residues were fixed. The optimal binding orientation
was achieved by utilizing 300 steps of steepest descent followed
Acknowledgements
We are grateful to the Canadian Institutes of Health
Research (MOP-14712) for financial support of this
research and to Rx&D-HRF/CIHR for a graduate
scholarship (to P.R.).
References and Notes
1. (a) Silverstein, F. E.; Faich, G.; Goldstein, J. L.; Simon,
L. S.; Pincus, T.; Whelton, A.; Makuch, R.; Eisen, G.; Agra-
wal, N. M.; Stenson, W. F.; Burr, A. M.; Zhao, W. W.; Kent,
J. D.; Lefkowith, J. B.; Verburg, K. M.; Geis, G. S. JAMA
2000, 284, 1247. (b) Hawkey, C.; Laine, L.; Simon, T.; Beau-
lieu, A.; Maldonado, C. J.; Acevedo, E.; Shahane, A.; Quan,
H.; Bolognese, J.; Mortensen, E. Arthritis Rheum. 2000, 43,
370.

P. N. P. Rao et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2205–S2209
2209
by the conjugate gradient method. The CVFF was employed
for all docking purposes. These docked structures were very
similar to the minimized structures obtained initially. The
quality of the docked structures were evaluated by measuring
the intermolecular energy of the ligand–enzyme assembly.
13. Analytical data for 12b. Mp 175–176 ꢀC; IR (KBr): 1718
d 1.46 (3H, t, J=7.3 Hz, SCH2CH3), 3.03 (3H, s, SO2CH3),
3.15 (2H, q, J=7.3 Hz, SCH2CH3), 6.35 (1H, s, pyranone
H-5), 7.04–7.07 (2H, m, phenyl H-2, H-6), 7.21–7.31 (3H, m,
phenyl H-3, H-4, H-5), 7.35 (2H, d, J=8.5 Hz, 4-MeSO2–
C6H4– H-2, H-6), 7.78 (2H, d, J=8.5 Hz, MeSO2–C6H4– H-3,
H-5). Anal. calcd for C20H18O4S2: C 62.16, H 4.69. Found: C
62.04, H 4.85.
1
(C¼O), 1314, 1153 (SO2) cmꢁ1; H NMR (300 MHz, CDCl3):