Synthesis and biological evaluation of 1-(benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene regioisomers: A novel class of 5-lipoxygenase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2008.05.071

A hitherto unknown class of linear acetylene regioisomers were designed such that a SO₂NH₂ group was located at the ortho-, meta-, or para-position of the acetylene C-1 phenyl ring, and a N-hydroxypyridin-2(1H)-one moiety was attached via its C-5 position to the C-2 position on an acetylene template (scaffold). All three regioisomers inhibited 5-lipoxygenase (5-LOX), where the relative potency order was 2-SO₂NH₂ (IC₅₀ = 10 μM) >3-SO₂NH₂ (IC₅₀ = 15 μM) >4-SO₂NH₂ (IC₅₀ = 68 μM) relative to the reference drug nordihydroguaiaretic acid (NDGA; IC₅₀ = 35 μM). The 2-SO₂NH₂ regioisomer (ED₅₀ = 86.0 mg/kg po) exhibited excellent oral anti-inflammatory (AI) activity that was more potent than aspirin (ED₅₀ = 128.9 mg/kg) and marginally less potent than ibuprofen (ED₅₀ = 67.4 mg/kg). The N-hydroxypyridin-2(1H)one moiety provides a novel pharmacophore for the design of cyclic hydroxamic mimetics capable of chelating 5-LOX iron for exploitation in the design of 5-LOX inhibitory AI drugs.

Full text of DOI:10.1016/j.bmcl.2008.05.071

Bioorganic & Medicinal Chemistry Letters 18 (2008) 4195–4198
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Bioorganic & Medicinal Chemistry Letters
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Synthesis and biological evaluation of 1-(benzenesulfonamido)-
2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene regioisomers:
A novel class of 5-lipoxygenase inhibitors
Morshed Alam Chowdhury a, Hua Chen b, Khaled R. A. Abdellatif a, Ying Dong a, Kenneth C. Petruk b,
Edward E. Knaus a,
*
a Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alta., Canada T6G 2N8
b Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alta., Canada T6G 2R7
a r t i c l e i n f o
a b s t r a c t
Article history:
A hitherto unknown class of linear acetylene regioisomers were designed such that a SO2NH2 group was
located at the ortho-, meta-, or para-position of the acetylene C-1 phenyl ring, and a N-hydroxypyridin-
2(1H)-one moiety was attached via its C-5 position to the C-2 position on an acetylene template
(scaffold). All three regioisomers inhibited 5-lipoxygenase (5-LOX), where the relative potency order
Received 4 April 2008
Revised 16 May 2008
Accepted 16 May 2008
Available online 22 May 2008
was 2-SO2NH2 (IC50 = 10
l
M) >3-SO2NH2 (IC50 = 15
l
M) >4-SO2NH2 (IC50 = 68
lM) relative to the refer-
ence drug nordihydroguaiaretic acid (NDGA; IC50 = 35
l
M). The 2-SO2NH2 regioisomer (ED50 = 86.0 mg/
Keywords:
Linear acetylenes
N-hydroxypyridin-2(1H)-ones
Lipoxygenase inhibition
Anti-inflammatory activity
kg po) exhibited excellent oral anti-inflammatory (AI) activity that was more potent than aspirin
(ED50 = 128.9 mg/kg) and marginally less potent than ibuprofen (ED50 = 67.4 mg/kg). The N-hydroxypyri-
din-2(1H)one moiety provides a novel pharmacophore for the design of cyclic hydroxamic mimetics
capable of chelating 5-LOX iron for exploitation in the design of 5-LOX inhibitory AI drugs.
Ó 2008 Elsevier Ltd. All rights reserved.
The most successful effort to develop 5-lipoxygenase (5-LOX)
inhibitors encompasses hydroxamic acids and related N-hydroxyu-
reas that likely chelate iron present in the 5-LOX enzyme.1 Some
representative examples of iron-chelating 5-LOX inhibitors (1,2
2,1 33) are shown in Figure 1. In a recent study, we described a
group of 1-aryl-2-(pyridyl)acetylene regioisomers (4) that consti-
tute a suitable scaffold (template) to design novel acyclic inhibitors
of the 5-LOX isozyme.4 It was anticipated that replacement of the
pyridyl ring in the acetylenes 4 by a N-hydroxypyridin-2(1H)one
moiety, that has the potential to chelate iron, may provide a hith-
erto unknown class of 5-LOX inhibitory anti-inflammatory agents.
Accordingly, we now describe the synthesis of a novel exploratory
group of 1-(2-, 3-, and 4-benzenesulfonamido)-2-[5-(N-hydroxy-
pyridin-2(1H)-one)]acetylene regioisomers (12a–c), their in vitro
evaluation as 5-LOX inhibitors, and in vivo assessment as anti-
inflammatory (AI) agents.
HO
NH2
O
Me
N
MeO
Cl
N
N
N
OH
S
O
Tepoxalin (2)
m-
Zileuton (1)
O
o-
SO2NH2
p-
Me
N
Het
C C
OH
3
4, Het = 2-, 3-, 4-pyridyl
Figure 1. Some representative iron-chelating 5-LOX inhibitors (1–3).
methodologies shown in Scheme 1. A modified procedure5 was
used to synthesize 5-ethynyl-2-methoxypyridine (8). Thus, Sono-
gashira coupling of 5-bromo-2-methoxypyridine (5) with 2-meth-
ylbut-3-yn-2-ol (6) in the presence of Et3N, CuI, and PdCl2(PPh3)2
catalyst afforded 4-(2-methoxypyridin-5-yl)-2-methyl-but-3-yn-
2-ol (7) in 75% yield. Subsequent removal of the isopropanol moi-
ety using sodium hydride furnished 5-ethynyl-2-methoxypyridine
(8) in 82% yield. A second Sonogashira cross-coupling reaction of 8
with the bromobenzenesulfonamide (9a–c) regioisomers was
carried out under an argon atmosphere in Et3N-THF (1:1, v/v) using
PdCl2(PPh3)2/CuI as catalyst to furnish the respective 1-(benzene-
sulfonamido)-2-(2-methoxypyrid-5-yl)acetylenes 10a–c in 26–72%
The
1-(benzenesulfonamido)-2-(2-methoxypyrid-5-yl)acety-
lenes (10a–c) were prepared using two consecutive palladium-cat-
alyzed Sonogashira cross-coupling reactions. The subsequent
transformation of 10a–c to the target 1-(2-, 3, and 4-benzenesul-
fonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene regioi-
somers (12a–c) was carried out using the synthetic
* Corresponding author. Tel.: +1 780 492 5993; fax: +1 780 492 1217.
E-mail address: eknaus@pharmacy.ualberta.ca (E.E. Knaus).
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.05.071

4196
M. A. Chowdhury et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4195–4198
Br
Me
Me
Me
a
H
C
C
OH
MeO
C
C
OH
MeO
N
5
N
Me
6
7
b
c
R1
MeO
C
C H
4
N
3
2
R2
8
9a, R1 = SO2NH2, R2 = 2-Br
9b, R1 = SO2NH2, R2 = 3-Br
9c, R1 = SO2NH2, R2 = 4-Br
2
3
R1
4
R1
d
MeO
C
C
MeO
C
C
+
N
N
10
11
O
R1
10-12: a, R1 = 2-SO2NH2
b, R1 = 3-SO2NH2
e
O
C
C
N
c, R1 = 4-SO2NH2
HO
12
Scheme 1. Reagents and conditions: (a) Et3N, Pd(PPh3)2Cl2, CuI, 70–75 °C, 3 h; (b) benzene, NaH, 105–110 °C, 1 h; (c) Et3N/THF, Pd(PPh3)2Cl2, CuI, 90 °C, 5 h;
(d) m-chloroperoxybenzoic acid, CH2Cl2, 25 °C, overnight; (e) i—acetyl chloride, reflux, 1 h, ii—MeOH, 25 °C, overnight.
yield. Oxidation of 10a–c with meta-chloroperbenzoic acid in
dichloromethane6 afforded the N-oxides 11a–c in 22–42% yield. Fi-
nally, reaction of 11a–c with acetyl chloride at reflux, and then
methanolysis in place of hydrolysis,6 furnished the target N-
hydroxypyridin-2(1H)-ones (12a–c) in 33–94% yield.
Replacement of the carboxyl (CO2H) in traditional non-steroidal
anti-inflammatory drugs (NSAIDs) by a hydroxamic acid (CON-
HOH) moiety provided potent orally active 5-LOX inhibitory
agents.3 NSAIDs having a CONHOH or CON(Me)OH (pKa 9–11
range) in place of the CO2H in traditional NSAIDs (pKa generally
in the 4–5 range) are much less acidic which decreases
ulcerogenicity.7
The rational for the design of the acyclic acetylenes 12a–c is
based on the expectations that the N-hydroxypyridin-2(1H)one
moiety will confer 5-LOX inhibitory activity. The CONOH part of
the N-hydroxypyrid-2(1H)-one ring present in 12a–c can be
viewed as a cyclic hydroxamic acid mimetic. These N-hydroxypyri-
din-2(1H)-ones, like acyclic hydroxamic acids, are expected to
serve as effective iron chelators to exhibit 5-LOX inhibitory activ-
ity. However, these cyclic N-hydroxypyridin-2(1H)-ones, unlike
acyclic hydroxamic acids which undergo facile biotransformation
to the acids, are expected to be metabolically stable with improved
oral efficacy. Compounds that possess a N-hydroxypyridin-2(1H)-
one moiety, in view of their low acidity, are expected to be non-
ulcerogenic.
in Table 1). The structure-activity data acquired showed that the
active 2-SO2NH2 regioisomer 12a (ED50 = 86.0 mg/kg po), unlike
the inactive 3-SO2NH2 12b and 4-SO2NH2 12c regioisomers, exhib-
ited an AI activity that was more potent than the aspirin
(ED50 = 128.9 mg/kg po) and marginally less potent than ibuprofen
(ED50 = 67.4 mg/kg po). Both aspirin and ibuprofen are over-the-
counter (OTC) non-prescription drugs used extensively to treat
minor pain and inflammation.
In conclusion, a previously unknown class of 1-(2-, 3-, and 4-
benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acety-
lene regioisomers8 were designed as 5-LOX inhibitors of inflamma-
tion. The structure-activity data acquired indicate that (i) a linear
acetylene spacer between vicinal 2-benzenesulfonamide and
Table 1
In vitro cell-based 5-LOX inhibition data and anti-inflammatory (AI) activities, for
1-(benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylenes (12a–c)
2
3
R1
4
O
C
C
N
HO
12a-c
An in vitro cell-based inhibition assay was used to determine
the biological effect of compounds 12a–c on eicosanoid synthe-
sis/release by measuring the amounts of cysteinyl leukotrienes
(collectively referred to as a group of 5-LOX derived metabolites
LTC4, LTD4, and LTE4) secreted into the culture medium of human
brain cancer cells. In this 5-LOX inhibition assay, all three SO2NH2
regioisomers 12a–c inhibited 5-LOX, where the relative potency
Compound
R1
5-LOX inhibitory activity:
AI activity:b
ED50 (mg/kg)
IC50
(l
M)a
12a
12b
12c
Aspirin
Ibuprofen
NDGAc
2-SO2NH2
3-SO2NH2
4-SO2NH2
—
—
—
10
15
68
—
—
35
86.0
Inactive
Inactive
128.9
67.4
—
order was 2-SO2NH2 (12a, IC50 = 10
IC50 = 15 M) >4-SO2NH2 (12c, IC50 = 68
ence drug NDGA (IC50 = 35 M).
l
M) >3-SO2NH2 (12b,
a
Values are the mean of two determinations and the deviation from the mean is
<10% of the mean value.
l
lM) relative to the refer-
l
b
Anti-inflammatory activity in a carrageenan-induced rat paw edema assay. The
The oral AI activities (ED50 values) exhibited by the cyclic N-
hydroxypyridin-2(1H)-one regioisomers 12a–c were determined
using a carrageenan-induced rat foot paw edema model (see data
results are expressed as the ED50 value (mg/kg) at 3 h after oral (po) administration
of the test compound.
c
NDGA, nordihydroguaiaretic acid was used as the Ref. 5-LOX inhibitory drug.

M. A. Chowdhury et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4195–4198
4197
1140 (SO2) cmÀ1 1H NMR (CDCl3+DMSO-d6) d 3.87 (s, 3H, OMe), 6.74 (d,
;
N-hydroxypyridin-2(1H)-one rings constitutes a potential tem-
plate for the design of new acyclic 5-LOX inhibitors, (ii) the pres-
ence and position of a SO2NH2 substituent on the phenyl ring is a
determinant of 5-LOX inhibitory activity9 where the relative po-
tency profile is 2- >3- and 4-SO2NH2, (iii) the N-hydroxypyrid-
2(1H)-one moiety provides a new 5-LOX pharmacophore for the
design of cyclic hydroxamic mimetics and (iv) 1-(2-benzenesulfo-
namidol)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene (12a) that
exhibits excellent oral anti-inflammatory activity,10 relative to the
reference drugs aspirin and ibuprofen, is a useful lead-compound
to further develop the 5-LOX drug design strategy described
herein.
J = 8.5 Hz, 1H, pyridyl H-3), 7.20 (br s, 2H, SO2NH2 that exchanges with D2O),
7.56 (d, J = 8.5 Hz, 2H, phenyl H-2, H-6), 7.69 (dd, J = 8.5, 2.4 Hz, 1H, pyridyl H-
4), 7.82 (d, J = 8.5 Hz, 2H, phenyl H-3, H-5), 8.27 (d, J = 2.4 Hz, 1H, pyridyl H-6).
General procedure for the synthesis of 1-(benzenesulfonamido)-2-(1-oxido-2-
methoxypyrid-5-yl)acetylene (11a–c). m-Chloroperoxybenzoic acid (77% max.)
(12 mmol) was added to a stirred solution of a 1-(benzenesulfonamido)-2-(2-
methoxypyrid-5-yl)acetylene (10) (2 mmol) in dry CH2Cl2 (50 mL), and the
reaction was allowed to proceed with stirring at 25 °C overnight. The solvent
CH2Cl2 was removed in vacuo to give a crude product which was purified by
silica gel column chromatography using methanol-EtOAc (2:3, v/v) as eluent to
afford the respective products 11a–c.
1-(2-Benzenesulfonamido)-2-(1-oxido-2-methoxypyrid-5-yl)acetylene
(11a).
Yield, 22%; pale yellow solid; mp 190–192 °C, IR (film): 3150–2800 (broad)
(NH2), 2150 (CꢀC), 1325, 1165 (SO2) cmÀ1
;
1H NMR(CDCl3+DMSO-d6) d 4.08 (s,
3H, OMe), 7.26 (d, J = 8.9 Hz, 1H, pyridyl H-3), 7.29 (br s, 2H, SO2NH2 that
exchanges with D2O), 7.45–7.58 (m, 2H, pyridyl H-4, phenyl H-5), 7.60–7.69
(m, phenyl H-4, H-6), 7.95 (dd, J = 7.6, 1.2 Hz, 1H, phenyl H-3), 8.62 (d,
J = 1.8 Hz, 1H, pyridyl H-6).
Acknowledgment
1-(3-Benzenesulfonamido)-2-(1-oxido-2-methoxypyrid-5-yl)acetylene
(11b).
We are grateful to the Canadian Institutes of Health Research
(CIHR) (MOP-14712) for financial support of this research.
Yield, 41%; yellowish solid; mp 150–152 °C; IR (film): 3200–2700 (broad)
(NH2), 2150 (CꢀC), 1325, 1165 (SO2) cmÀ1
;
1H NMR (CDCl3+DMSO-d6) d 4.05 (s,
3H, OMe), 7.18 (d, J = 8.9 Hz, 1H, pyridyl H-3), 7.34 (br s, 2H, SO2NH2 that
exchanges with D2O), 7.40 (dd, J = 7.9, 7.6 Hz, 1H, phenyl H-5), 7.45–7.55 (m,
2H, pyridyl H-4, phenyl H-6), 7.66 (ddd, J = 7.9, 1.5, 1.5 Hz, 1H, phenyl H-4),
7.97 (dd, J = 1.5, 1.5 Hz, 1H, phenyl H-2), 8.40 (d, J = 1.8 Hz, 1H, pyridyl H-6).
References and notes
1-(4-Benzenesulfonamido)-2-(1-oxido-2-methoxypyrid-5-yl)acetylene
(11c).
1. Muri, E. M. F.; Nieto, M. J.; Sindelar, R. D.; Williamson, J. S. Curr. Med. Chem.
2002, 9, 1631.
Yield, 42%; white solid; mp 205–207 °C; IR (film): 3250–2700 (broad) (NH2),
2225 (CꢀC), 1365, 1160 (SO2) cmÀ1
;
1H NMR (CDCl3+DMSO-d6) d 4.05 (s, 3H,
2. Carter, G. W.; Young, P. R.; Albert, D. H.; Bouska, J.; Dyer, R.; Bell, R. L.;
Summers, J. B.; Brooks, D. W. J. Pharmacol. Exp. Ther. 1991, 256, 929.
3. Summers, J. B.; Gunn, B. P.; Mazdiyasni, H.; Goetze, A. M.; Young, P. R.; Bouska,
J. B.; Dyer, R. D.; Brooks, D. W.; Carter, G. W. J. Med. Chem. 1987, 30, 2121.
4. Chowdhury, M. A.; Dong, Y.; Chen, Q.-H.; Abdellatif, K. R. A.; Knaus, E. E. Bioorg.
Med. Chem. 2008, 16, 1948.
5. Capraro, H. -G.; Furet, P.; Garcia-Echeverria, C.; Stauffer, F. PCT Int. Appl. 2005,
WO 2005054238.
6. Choi, H.-Y.; Yoon, S.-H. Bull. Korean Chem. Soc. 1999, 20, 857.
7. U. Chiodoni. U.S. Patent 4,455,432, Issued June 19, 1984.
OMe), 7.23 (d, J = 8.9 Hz, 1H, pyridyl H-3), 7.37 (br s, 2H, SO2NH2 that exchanges
with D2O), 7.52 (dd, J = 8.9, 1.8 Hz, 1H, pyridyl H-4), 7.65 (d, J = 8.5 Hz, 2H,
phenyl H-2, H-6), 7.85 (d, J = 8.5 Hz, 2H, phenyl H-3, H-5), 8.42 (d, J = 1.8 Hz,
1H, pyridyl H-6).
General procedure for the synthesis of 1-(benzenesulfonamido)-2-[5-(N-
hydroxypyridin-2(1H)-one)]acetylenes (12a–c). Acetyl chloride (6 mL) was
added to 11a, 11b, or 11c (2 mmol) and the reaction was allowed to proceed
at reflux for 1 h. The reaction mixture was cooled to 25 °C, and excess acetyl
chloride was removed in vacuo. The residue was dissolved in methanol prior to
stirring at 25 °C overnight. Methanol was removed in vacuo to give a solid
product which was then mixed with Et2O (10 mL) to form a slurry. Finally, the
product was filtered out and dried under vacuum to give the respective
products (12a–c).
8. Experimental and spectral data for compounds 7–12. 4-(2-Methoxypyridin-5-
yl)-2-methylbut-3-yn-2-ol (7). PdCl2(PPh3)2 (63 mg, 0.09 mmol) and CuI (19 mg,
0.10 mmol) were added to a stirred solution of 5 (2.75 mL, 21.39 mmol) and 6
(2.20 mL, 22.61 mmol) in Et3N (40 mL) under an argon atmosphere at 25 °C,
and the reaction was allowed to proceed at 70–75 °C for 3 h. The reaction
mixture was allowed to cool to 25 °C, filtered, and excess Et3N was removed
from the filtrate in vacuo. The dark brown residue obtained was purified by
silica gel column chromatography using hexane-EtOAc (3:1, v/v) as eluent to
1-(2-Benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene (12a).
Yield, 94%; pale yellow solid; mp 221–223 °C (decomp.); IR (film): 3350–
2900 (broad) (NH2), 2200 (CꢀC), 1662 (CO), 1350, 1140 (SO2) cmÀ1
;
1H NMR
(CDCl3+DMSO-d6) d 6.14 (d, J = 9.8 Hz, 1H, pyridone H-3), 6.45 (br s, 2H, SO2NH2
that exchanges with D2O), 6.97 (d, J = 6.7, 6.7 Hz, 1H, phenyl H-5), 7.02–7.13
(m, 2H, pyridone H-4, phenyl H-4), 7.18 (d, J = 6.7 Hz, 1H, phenyl H-6), 7.56 (d,
J = 6.7 Hz, 1H, phenyl H-3), 7.74 (br s, 1H, pyridone H-6); 13C NMR (DMSO-d6) d
86.3 (acetylene C-2), 92.5 (acetylene C-1), 99.2 (pyridone C-5), 119.1 (pyridone
C-3), 119.7 (phenyl C-1), 127.2 (pyridone C-6), 128.3 (phenyl C-3), 131.7
(phenyl C-4), 133.6 (phenyl C-5), 139.9 (pyridone C-4), 140.0 (phenyl C-6),
144.0 (phenyl C-2), 157.0 (C@O); MS (ES+) m/z: Calcd for C13H10N2O4S (MH+):
291.04. Found: 290.84.
afford 7 in 75% yield; yellowish oil; IR (film): 3368 (OH), 2235 (CꢀC) cmÀ1
;
1H
NMR (CDCl3) d 1.62 (s, 6H, CMe2), 2.44 (br s, 1H, OH), 3.94 (s, 3H, OMe), 6.69 (d,
J = 8.5 Hz, 1H, pyridyl H-3), 7.59 (dd, J = 8.5, 2.1 Hz, 1H, pyridyl H-4), 8.27 (d,
J = 2.1 Hz, 1H, pyridyl H-6).
5-Ethynyl-2-methoxypyridine (8). Sodium hydride (26 mg, 1.08 mmol) was
added to a solution of 7 (1.52 g, 7.96 mmol) in benzene (7 mL), and the
reaction mixture was heated at 105–110 °C for 1 h. Removal of the solvent in
vacuo gave
a dark brown oil which was purified by silica gel column
1-(3-Benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene (12b).
Yield, 33%; yellowish solid; mp 180-182 °C (decomp.); IR (film): 3300–2850
chromatography using hexane-EtOAc (3:1, v/v) as eluent to afford 8 in 82%
yield; brown oil; IR (film): 2230 (CꢀC) cmÀ1
;
1H NMR (CDCl3) d 3.12 (s, 1H,
(broad) (NH2), 2200 (CꢀC), 1660 (CO), 1350, 1160 (SO2) cmÀ1
;
1H NMR
CꢀCH), 3.96 (s, 3H, OMe), 6.72 (d, J = 8.5 Hz, 1H, pyridyl H-3), 7.66 (dd, J = 8.5,
(CDCl3+DMSO-d6) d 6.51 (d, J = 8.9 Hz, 1H, pyridone H-3), 7.28 (br s, 2H, SO2NH2
that exchanges with D2O), 7.40 (dd, J = 8.9 Hz, 1H, pyridone H-4), 7.48 (dd,
J = 7.9, 7.6 Hz, 1H, phenyl H-5), 7.59 (d, J = 7.6 Hz, 1H, phenyl H-6), 7.79 (d,
J = 7.9 Hz, 1H, phenyl H-4), 7.90 (s, 1H, phenyl H-2), 8.09 (s, 1H, pyridone H-6),
12.05 (br s, 1H, N–OH); 13C NMR (DMSO-d6) d 86.7 (acetylene C-2), 87.6
(acetylene C-1), 98.7 (pyridone C-5), 119.2 (pyridone C-3), 122.9 (phenyl C-1),
125.4 (phenyl C-4), 127.9 (pyridone C-6), 129.5 (phenyl C-5), 133.9 (phenyl C-
2), 139.7 (phenyl C-6), 139.9 (pyridone C-4), 144.5 (phenyl C-3), 156.9 (C@O);
MS (ES+) m/z: Calcd for C13H10N2O4S (MH+): 291.04. Found: 290.84.
2.1 Hz, 1H, pyridyl H-4), 8.27 (d, J = 2.1 Hz, 1H, pyridyl H-6).
General procedure for the synthesis of 1-(benzenesulfonamido)-2-(2-
methoxypyrid-5-yl)acetylenes (10a–c). CuI (46 mg, 0.24 mmol) was added
with stirring to
a solution containing PdCl2(PPh3)2 (85 mg, 0.12 mmol), 8
(6 mmol), and a bromobenzenesulfonamide 9a, 9b, or 9c (4 mmol) in dry THF
(10 mL) and Et3N (10 mL) under an argon atmosphere. The reaction mixture
was heated at 90 °C for 5 h, cooled to 25 °C, and filtered to remove the
inorganic salts. The solvent from the filtrate was removed in vacuo, and the
residue obtained was purified by silica gel column chromatography using
hexane-acetone (1:1, v/v) as eluent to furnish the respective products 10a–c.
1-(2-Benzenesulfonamido)-2-(2-methoxypyrid-5-yl)acetylene (10a). The product
was obtained as a pale yellow oil using the Sonogashira coupling reaction of 8
with 9a in 26% yield; IR (film): 3380, 3250 (NH2), 2160 (CꢀC), 1295, 1150 (SO2)
1-(4-Benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene (12c).
Yield, 48%; dark brown solid; mp 125–127 °C; IR (film): 3150–2900 (board)
(NH2), 2180 (CꢀC), 1650 (C@O), 1340, 1140 (SO2) cmÀ1
;
1H NMR
(CDCl3+DMSO-d6) d 6.23 (d, J = 9.2 Hz, 1H, pyridone H-3), 6.65 (br s, 2H,
SO2NH2 that exchanges with D2O), 7.06 (d, J = 9.2 Hz, 1H, pyridone H-4), 7.18
(d, J = 7.6 Hz, 2H, phenyl H-2, H-6), 7.51 (d, J = 7.6 Hz, 2H, phenyl H-3, H-5),
7.60 (s, 1H, pyridone H-6), 11.55 (br s, 1H, N–OH); 13C NMR (DMSO-d6) d 87.8
(acetylene C-2), 88.0 (acetylene C-1), 98.7 (pyridone C-5), 119.2 (pyridone C-3),
125.6(phenyl C-1), 126.0 (phenyl C-2, C-6), 127.8 (pyridone C-6), 131.4 (phenyl
C-3, C-5), 140.0 (pyridone C-4), 143.5 (phenyl C-4), 156.9 (C@O); MS (ES+) m/z:
Calcd for C13H10N2O4S (MH+): 291.04. Found: 290.84; Anal. Calcd for
cmÀ1 1H NMR (CDCl3) d 3.97 (s, 3H, OMe), 5.28 (br s,2H, SO2NH2), 6.75 (d,
;
J = 8.5 Hz, 1H, pyridyl H-3), 7.45 (ddd, J = 7.6, 7.6, 1.2 Hz, 1H, phenyl H-5), 7.55
(ddd, J = 7.6, 7.6, 1.2 Hz, 1H, phenyl H-4), 7.72 (dd, J = 7.6, 1.2 Hz, 1H, phenyl H-
6), 7.77 (dd, J = 8.5, 2.1 Hz, 1H, pyridyl H-4), 8.05 (dd, J = 7.6, 1.2 Hz, 1H, phenyl
H-3), 8.41 (d, J = 2.1 Hz, 1H, pyridyl H-6).
1-(3-Benzenesulfonamido)-2-(2-methoxypyrid-5-yl)acetylene (10b). The product
was obtained as a dark oil using the Sonogashira coupling reaction of 8 with 9b
in 72% yield; IR (film): 3360, 3270 (NH2), 2200 (CꢀC), 1315, 1150 (SO2) cmÀ1
;
C
13H10N2O4S Á H2O: C, 50.65; H, 3.92. Found: C, 50.79; H, 3.96.
1H NMR (CDCl3+DMSO-d6) d 3.98 (s, 3H, OMe), 5.08 (br s, 2H, SO2NH2 that
exchanges with D2O), 6.76 (d, J = 8.6 Hz, 1H, pyridyl H-3), 7.51 (dd, J = 7.9,
7.6 Hz, 1H, phenyl H-5), 7.65-7.72 (m, 2H, pyridyl H-4, phenyl H-6), 7.87 (ddd,
J = 7.9, 1.5, 1.5 Hz, 1H, phenyl H-4), 8.08 (dd, J = 1.5, 1.5 Hz, 1H, phenyl H-2),
8.36 (d, J = 2.1 Hz, 1H, pyridyl H-6).
9. In vitro cell-based enzyme immunoassay for determination of cysteinyl
leukotrienes (5-LOX). The biological effects of the test compounds 12a–c on
eicosanoid synthesis/release were determined by measuring the amounts of
cysteinyl leukotrienes (collectively referred to as a group of 5-LOX derived
metabolites LTC4, LTD4, and LTE4) secreted into the culture medium of
human brain cancer cells. Primary culture of ED 273b-BT human
glioblastoma cells derived from patient was established and characterized
in our laboratory as previously described (Farr-Jones, M. A.; Parney, I. F.;
1-(4-Benzenesulfonamido)-2-(2-methoxypyrid-5-yl)acetylene (10c). The product
was obtained as a white solid using the Sonogashira coupling reaction of 8 with
9c in 70% yield; mp 185–187 °C; IR (film): 3340, 3250 (NH2), 2190 (CꢀC), 1290,

4198
M. A. Chowdhury et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4195–4198
Petruk, K. C. J. Neurooncology 1999, 43, 399). Cells were seeded in 12-well
2000 rpm, and stored at À80 °C until assayed. The concentrations of
eicosanoids were determined using a cysteinyl leukotriene (catalog number
520501) enzyme immunoassay kit (Cayman Chemical, Ann Arbor, MI, USA)
according to a previously reported method (Uddin, M. J.; Rao, P. N. P.; Knaus,
E. E. Bioorg. Med. Chem. 2004, 12, 5929).
plates (2 Â 105cells/well) and cultured in Dulbecco’s modified Eagle’s
medium and F-12 nutrition mixture (Invitrogen, Grand Islands, NY, USA)
supplemented with 10% heat-inactivated fetal calf serum, 100 U/mL
penicillin, and 100 units/mL streptomycin at 37 °C in
a humidified
atmosphere of 5% CO2. The cells were stimulated with the appropriate
concentrations of the test compounds and the positive control NDGA
10. In vivo anti-inflammatory assay. The test compounds 12a–c and the reference
drugs aspirin and ibuprofen were evaluated using the in vivo carrageenan-
induced foot paw edema model reported previously (Winter, C. A.; Risley, E. A.;
Nuss, G. W. Proc. Soc. Exp. Biol. Med. 1962, 111, 544).
(Cayman Chemical, Ann Arbor, MI, USA) for 5-LOX activity. After
a 24 h
incubation, supernatants were harvested, centrifuged for 10 min at