Discovery and synthesis of a potent sulfonamide ET(B) selective antagonist

Article abstract of DOI:10.1016/S0960-894X(00)00366-8

The synthesis and structure-activity relationships of a series of sulfonamide endothelin antagonists are described. In the course of our modification studies, we discovered ET(B) selective antagonists. The most potent compound 15f displays IC₅₀ values of 1.7 μM and 0.002 μM to ET(A) and ET(B) receptors, respectively. (C) 2000 Elsevier Science Ltd. All rights reserved.

Full text of DOI:10.1016/S0960-894X(00)00366-8

Bioorganic & Medicinal Chemistry Letters 10 (2000) 1875±1878
Discovery and Synthesis of a Potent Sulfonamide ETB
Selective Antagonist
Yasuhiko Kanda,a,* Tadashi Takahashi,a Yoshitaka Araki,a Toshiro Konoike,b,*
Shin-ichi Miharaa and Masafumi Fujimotoa
aShionogi Research Laboratories, Shionogi & Co., Ltd., Fukushima-ku, Osaka 553-0002, Japan
bShionogi Research Laboratories, Shionogi & Co., Ltd., Amagasaki, Hyogo 660-0813, Japan
Received 1 May 2000; accepted 21 June 2000
AbstractÐThe synthesis and structure±activity relationships of a series of sulfonamide endothelin antagonists are described. In the
course of our modi®cation studies, we discovered ETB selective antagonists. The most potent compound 15f displays IC50 values of
1.7 mM and 0.002 mM to ETA and ETB receptors, respectively. # 2000 Elsevier Science Ltd. All rights reserved.
Introduction
ETA selective antagonists. In this paper, we wish to
describe the discovery and synthesis of a potent ETB
selective antagonist.
Endothelins (ET-1, ET-2, ET-3), 21-amino acid bicyclic
peptides, are the most potent known vasoconstrictors.1,2
Two distinct G-protein coupled receptors, ETA and
ETB, were cloned and characterized with respect to their
anity for each endothelin.3,4 The ETA receptor is
expressed on vascular smooth muscle cells and has high
anity for ET-1 and ET-2. The ETB receptor is expres-
sed on vascular endothelial and smooth muscle cells and
has high anity for all three endothelins. As these two
subtypes of receptors are widely distributed in human
tissues, the development of selective or non-selective
endothelin antagonists is expected to be useful for the
treatment of various diseases.5 A number of groups
have reported the discovery of non-peptide endothelin
antagonists since 1994. Most of them were ETA selective
or non-selective antagonists,6,7 and only recently have
non-peptide ETB selective antagonists been reported.8
Figure 1.
Chemistry
Sulfonamides 5a±m of 4-substituted azoles (Table 1)
were synthesized by condensation of the substituted
benzenesulfonyl chlorides with the 4-substituted amino-
isoxazoles 4A (X=N, Y=O), 4B (X=O, Y=N) or 4-
substituted aminoisothiazoles 4C (X=N, Y=S), and
4D (X=S, Y=N) (Scheme 1). As most of the azoles 4
were not readily available, we developed a general syn-
thetic method for preparing 4-substituted aminoazoles 4
starting from 1.10 Our method was based on directed
ortho-metallation and was successfully used to prepare
4-substituted aminoisoxazoles 4A bearing various sub-
stituents (R2) at the 4-position. Substituted 4-lithioisox-
azole was prepared from 1A10 using alkyllithium for the
directed ortho-metallation, which was reacted with various
electrophiles (iodomethane, iodoethane, benzyl bromide,
methyl disul®de, disul®de 9) to give 3A. Deprotection of
the amino group gave 4A, from which sulfonamides
In order to ®nd low-molecular-weight non-peptide
endothelin antagonists, we screened the Shionogi com-
pound library for compounds capable of inhibiting spe-
ci®c [125I]ET-1 binding. We identi®ed sulfamethoxazole
5a9 and its iodide 5b as ETA selective antagonists (Fig.
1). We then started the modi®cation of 5b as a lead
compound to enhance the binding anity for the ETA
receptor. Our modi®cation study led to the discovery of
ETB selective antagonists as well as non-selective and
*Corresponding authors. Tel.: +81-6-6458-5861; fax: +81-6-6458-
0987; e-mail: yasuhiko.kanda@shionogi.co.jp, toshiro.konoike@
shionogi.co.jp
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00366-8

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Y. Kanda et al. / Bioorg. Med. Chem. Lett. 10 (2000) 1875±1878
Table 1.
carboxylic acid 15g as well as to ketone 15j and oxime
15l (Scheme 3).
Results and Discussion
IC50 (mM)
Structure±activity relationships were discussed using
IC50 values obtained from radioreceptor binding studies.
For ETA receptor binding assay, we employed rat aortic
smooth muscle A7r5 cells. For ETB receptor binding
assay, we employed COS-7 cells transfected with porcine
ETB receptor. IC50 data were recorded by measuring the
displacement of [125I]ET-1 binding from ETA receptor
or [125I]ET-3 binding from ETB receptor.
Compound
X
Y
R1
R2
ETA
ETB
5a
5b
5c
5d
5e
5f
5g
5h
5i
N
N
N
N
N
N
O
N
S
N
N
N
N
O
O
O
O
O
O
N
S
N
O
O
O
O
NH2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
t-Bu
t-Bu
t-Bu
t-Bu
t-Bu
H
I
Cl
Me
Et
PhCH2
Me
Me
333
11
3.5
5.0
3.4
71
9.0
28
ND
>1000
ND
ND
ND
ND
ND
ND
ND
30
Table 1 shows the in¯uences of substituent R2 in the
azole ring and substituent R1 in the sulfonamide of
compounds 5a±m. We expected that introduction of a
variety of groups for R2 would lead to the enhancement
of the binding anity for the ETA receptor because our
lead compound 5b had a higher binding anity than the
parent 5a. We ®rst investigated the in¯uence of the R2
substituent on the antagonistic activity. Halogen (5b,
5c) or small alkyl groups (5d, 5e) were needed for the
activity, but their binding anities were moderate. A
large benzyl substituent decreased the activity (5f). Iso-
meric isoxazole ring (5g) or isothiazole analogues (5h,
5i) had a lower binding anity than the corresponding
compound 5d.
Me
I
Me
>500
>100
28
5j
5k
5l
ND
35
MeS
3-MeO-PhS
>100
5m
0.53
0.30
ND, not determined.
5d±f, 5k±m were prepared by treatment with 4-acet-
amido- and 4-tert-butyl-benzenesulfonyl chlorides. In
contrast to the isoxazoles, we could not prepare iso-
thiazole derivatives (5h, 5i) eciently by our method
because a directed ortho-lithiation of 1C or 1D11 com-
peted with lithiation of the methyl group. Thus, 4-
lithioisothiazoles were prepared via halogen±metal
exchange reaction of bromide 2C or iodide 2D obtained by
halogenation of the parent aminoisothiazole. The resulting
lithium compounds were reacted with iodomethane to
give methylated isothiazoles, which were then converted
to sulfonamide 5h, 5i.
We next replaced the hydrophilic amino group R1 with
the hydrophobic tert-butyl group, and investigated the
e€ect of R2. Although compound 5j±l showed no or
decreased binding anity for the ETA receptor, com-
pounds 5j and 5l were found to be ETB selective
antagonists with moderate anity. This indicated that
introduction of a bulky hydrophobic substituent R1 is
e€ective for ETB anity. Therefore, in order to increase
the binding anity for both subtypes, we modi®ed the
4-position and introduced the 3-methoxy phenylthio
group by an analogy to the known non-selective endo-
thelin antagonist.7a By this modi®cation, we obtained
the non-selective antagonist 5m with improved IC50
values in the sub-micromolar range for both subtypes.
As a typical example of 15a±n, the synthesis of 15f is
shown in Scheme 2. The isoxazole ring was constructed
by 1,3-dipolar cycloaddition reaction of acetylene 7 and
the nitrile oxide which was prepared from 10.12 Hydro-
lysis of the ester gave carboxylic acid 11 which was
converted to the protected amine 12 by Curtius rear-
rangement. The 4-position of 12 was lithiated, and the
resulting lithium compound was treated with disul®de
913 to give 13. Silyloxy compound 13 was converted to
an acetate, and deprotection of its Boc group gave
amine 14. Sulfonylation of 14 with 4-tert-butyl-
benzenesulfonyl chloride, and subsequent hydrolysis of
the acetate gave sulfonamide 15d. Alcohol 15d was
converted to nitrile 15k, and reduction of the nitrile by
DIBAH gave aldehyde 15f. Aldehyde 15f was subse-
quently converted to ester 15h and amide 15i through
Next, we selected 5m as the second lead compound for a
non-selective endothelin antagonist, and modi®ed the 5-
position of the isoxazole ring (Table 2). One carbon
homologation of 5m decreased the anity for both
subtypes (15a). Therefore, we decided to introduce a
functional group with the expectation of speci®c interaction
Scheme 1. (a) n-BuLi, THF, 78 ꢀC to rt; (b) electrophile, THF, 78 ꢀC; (c) Br2, AcONa, AcOH, rt; (d) ICl, AcOH, rt; (e) NaH, THF, rt; (f) t-BuLi,
THF, 78 ꢀC; (g) MeI, THF, 78 ꢀC; (h) TFA, rt; (i) 4-AcNHPhSO2Cl, pyridine, rt; (j) aq NaOH, MeOH, re¯ux; (k) 4-t-BuPhSO2Cl, pyridine, rt.

Y. Kanda et al. / Bioorg. Med. Chem. Lett. 10 (2000) 1875±1878
1877
Scheme 2. (a) t-BuSiMe2Cl, imidazole, CH2Cl2, rt, 97%; (b) DMSO, 85 ꢀC, 80%; (c) Et3N, 7, rt, 76%; (d) aq NaOH, MeOH, rt, 81%; (e) SOCl2,
pyridine, CH2Cl2, 0 ꢀC; (f) NaN3, acetone±H2O, 0 ꢀC; (g) t-BuOH, toluene, re¯ux, 41% (three steps); (h) n-BuLi, THF, 78 ꢀC to rt; (i) 9, THF,
78 ꢀC, 67% (two steps); (j) n-Bu4NF, THF, rt; (k) Ac2O, DMAP, pyridine, CH2Cl2, rt; (l) TFA, anisole, rt, 86% (three steps); (m) 4-t-BuPhSO2Cl,
DMAP, pyridine, 50 ꢀC, 64%; (n) aq NaOH, MeOH±THF, rt, 92%; (o) MsCl, pyridine, 0 ꢀC; (p) aq NaOH, MeOH±THF, rt; (q) NaCN, DMF,
80 ꢀC; (q) DIBAH, toluene, 78 ꢀC to rt, 75% (four steps).
Table 2.
IC50 (mM)
ETB selectivity
(IC50 ETA/IC50 ETB)
Compound
n
R
ETA
ETB
5m
1
2
1
2
3
4
3
3
3
3
3
3
3
2
4
H
H
OH
OH
OH
OH
CHO
CO2H
CO2Me
CONH2
COCH3
CN
0.53 0.30
0.80 2.8
1.8
0.3
11
1.8
2.3
2.6
850
8.6
1.3
26
3.3
2.5
18
0.8
70
15a
15b
15c
15d
15e
15f
15g
15h
15i
15j
15k
15l
15m
15n
6.2
0.56
0.77 0.43
0.32 0.14
0.63 0.24
1.7
4.3
0.002
0.50
0.40 0.31
6.3
1.3
1.3
0.24
0.40
0.52
0.085
Scheme 3. (a) PDC, DMF, rt, 79%; (b) SOCl2, pyridine, Et2O, 0 ꢀC;
(c) MeOH, 0 ꢀC, 37% (two steps); (d) 28% aq NH3, 0 ꢀC, 24%; (e)
MeLi, THF, 78 ꢀC to 0 ꢀC; (f) PDC, DMF, rt, 29% (two steps); (g)
NH2OH HCl, EtOH±pyridine, rt, 100%.
CHˆNOH 1.5
CHO
CHO
0.38 0.50
1.4 0.020
between the functional group and the receptor. We ®rst
introduced a hydroxyalkyl group ((CH2)nOH) which
had both hydrogen bond donating and accepting prop-
erties and optimized the length of the alkyl chain (n=1±
4, 15b±e). Of these compounds, 15d (n=3) showed the
best anity for both subtypes. Next, we introduced
other functional groups, keeping the length of the alkyl
chain unchanged (n=3, 15f±l). Interestingly, aldehyde
15f had highly improved anity and selectivity for the
ETB receptor. The IC50 values of 15f were 1.7 mM and
0.002 mM for ETA and ETB receptors, respectively, and
the ETB selectivity (IC50 ETA/IC50 ETB) was 850. This
compound was found to be one of the most potent ETB
selective antagonists reported so far.8 Amide 15i and
oxime 15l were also ETB selective antagonists, although
they had a lower activity than 15f. Other derivatives,
such as carboxylic acid 15g, ester 15h, ketone 15j, and
nitrile 15k had decreased binding anity for both sub-
types compared with 15d, and their ETB selectivities
were low. Aldehyde analogues with shorter (n=2, 15m)

1878
Y. Kanda et al. / Bioorg. Med. Chem. Lett. 10 (2000) 1875±1878
or longer (n=4, 15n) alkyl spacers had lower activity
and selectivity for the ETB receptor than 15f. These
®ndings demonstrate that the substituent in the 5-
position of the isoxazole ring plays a crucial role in the
ETB receptor binding. Adequate spatial arrangement of
the speci®c functional group is needed for the high anity
binding to the ETB receptor. Our results indicate that
compound 15f displays speci®c interaction between its
aldehyde group and the ETB receptor binding site.
Wu, C.; Chan, M. F.; Stavros, F.; Raju, B.; Okun, I.; Castillo,
R. S. J. Med. Chem. 1997, 40, 1682±1689.
7. (a) Clozel, M.; Breu, V.; Gray, G. A.; Kalina, B.; Lo‚er, B.
M.; Burri, K.; Cassal, J. M.; Hirth, G.; Muuler, M.; Neidhart,
W.; Ramuz, H. J. Pharmacol. Exp. Ther. 1994, 270, 228±235.
(b) Elliott, J. D.; Lago, M. A.; Cousins, R. D.; Gao, A.; Leber,
J. D.; Erhard, K. F.; Nambi, P.; Elshourbagy, N. A.; Kumar,
C.; Lee, J. A.; Bean, J. W.; DeBrosse, C. W.; Eggleston, D. S.;
Brooks, D. P.; Feuerstein, G.; Ru€olo, R. R. J.; Weinstock, J.;
Gleason, J. G.; Peisho€, C. E.; Ohlstein, E. H. J. Med. Chem.
1994, 37, 1553±1557. (c) Walsh, T. F.; Fitch, K. J.; Chakravarty,
K.; Williams, D. L.; Murphy, K. A.; Nolan, N. A.; O'Brien, J.
A.; Lis, E. V.; Pettibone, D. J.; Kivlighn, S. D.; Gabel, R. A.;
Zingaro, G. J.; Krause, S. M.; Siegl, P. K. S.; Clineschmidt, B.
V.; Greenlee, W. J. 208th ACS National Meeting, Washington,
DC, August 1994; Medi 145. (d) Jae, H. S.; Winn, M.; Dixon, D.
B.; Marsh, K. C.; Nguyen, B.; Opgenorth, T. J.; von Geldern, T.
W. J. Med. Chem. 1997, 40, 3217±3227.
In conclusion, we have discovered a potent ETB selective
antagonist 15f. Unfortunately, 15f had insucient oral
bioavailability. However, it should be useful for under-
standing the role of the ETB receptor in pathological
conditions. Further modi®cation of 15f is in progress
and will be reported elsewhere.
8. (a) Chan, M. F.; Kois, A.; Verner, E. J.; Raju, B. G.; Cas-
tillo, R. S.; Wu, C.; Okun, I.; Stavros, F. D.; Balaji, V. N.
Bioorg. Med. Chem. 1998, 6, 2301±2316. (b) Mederski, W. W.
K. R.; Osswald, M.; Dorsch, D.; Christadler, M.; Schmitges,
C.-J.; Wilm, C. Bioorg. Med. Chem. Lett. 1999, 9, 619±622. (c)
von Geldern, T. W.; Tasker, A. S.; Sorensen, B. K.; Winn, M.;
Szczepankiewicz, B. G.; Dixon, D. B.; Chiou, W. J.; Wang, L.;
Wessale, J. L.; Adler, A.; Marsh, K. C.; Nguyen, B.; Opgen-
orth, T. J. J. Med. Chem. 1999, 42, 3668±3678. (d) Liu, G.;
Kozmina, N. S.; Winn, M.; von Geldern, T. W.; Chiou, W. J.;
Dixon, D. B.; Nguyen, B.; Marsh, K. C.; Opgenorth, T. J. J.
Med. Chem. 1999, 42, 3679±3689.
References and Notes
1. Yanagisawa, M.; Kurihara, H.; Kimura, S.; Tomobe, Y.;
Kobayashi, M.; Yazaki, Y.; Goto, K.; Masaki, T. Nature
1988, 332, 411±445.
2. Inoue, A.; Yanagisawa, M.; Kimura, S.; Kasuya, Y.;
Miyauchi, T.; Goto, K.; Masaki, T. Proc. Acad. Sci. USA
1989, 86, 2863±2867.
3. Arai, H.; Hori, S.; Aramori, I.; Ohkubo, H.; Nakanishi, S.
Nature 1990, 348, 730±732.
4. Sakurai, T.; Yanagisawa, M.; Takuwa, Y.; Miyazaki, Y.;
Kimura, S.; Goto, K.; Masaki, T. Nature 1990, 348, 732±735.
5. Webb, M. L.; Meek, T. D. Med. Res. Rev. 1997, 17, 17±67.
6. (a) Stein, P. D.; Hunt, J. T.; Floyd, D. M.; Moreland, S.;
Dickinson, K. E. J.; Mitchell, C.; Liu, E. C.-K.; Webb, M. L.;
Murugesan, N.; Dickey, J.; McMullen, D.; Zhang, R.; Lee, V.
G.; Sera®no, R.; Delaney, C.; Schae€er, T. R.; Kozlowski, M.
J. Med. Chem. 1994, 37, 329±331. (b) Doherty, A. M.; Patt,
W. C.; Edmunds, J. J.; Berryman, K.; Reisdoph, B. R.; Plum-
mer, M.; Shahripour, A.; Lee, C.; Cheng, X.-M.; Walker, D.
M.; Haleen, S.; Keiser, J. A.; Flynn, M. A.; Welch, K. M.;
Hallak, H. O.; Taylor, D. G.; Reynolds, E. E. J. Med. Chem.
1995, 38, 1259±1263. (c) Winn, M.; von Geldern, T. W.;
Opgenorth, T. J.; Jae, H.-S.; Tasker, A. S.; Boyd, S. A.;
Kester, J. A.; Mantei, R. A.; Bal, R.; Sorensen, B. K.;
Wu-Wong, J. R.; Chiou, W. J.; Dixon, D. B.; Novosad, E. I.;
Hernandes, L.; Marsh, K. C. J. Med. Chem. 1996, 39, 1039±
1048. (d) Riechers, H.; Albrecht, H. P.; Amberg, W.; Bau-
mann, E.; Bernard, H.; Bohm, H. J.; Klinge, D.; Kling, A.;
Muller, S.; Rashack, M.; Unger, L.; Walker, N.; Wernrt, W. J.
Med. Chem. 1996, 39, 2123±2128. (e) Sakurawi, K.; Yasuda,
F.; Tozyo, T.; Nakamura, M.; Sato, T.; Kikuchi, J.; Terui, Y.;
Ikenishi, Y.; Iwata, T.; Takahashi, K.; Konoike, T.; Mihara,
S.; Fujimoto, M. Chem. Pharm. Bull. 1996, 44, 343±351. (f)
9. Kano, H.; Ogata, K. Ann. Rep. Shionogi Res. Lab. 1957, 7,
1. See C.A. 1957, 51, 17889.
10. Konoike, T.; Kanda, Y.; Araki, Y. Tetrahedron Lett.
1996, 37, 3339±3342.
11. 5-(Boc-amino)-3-methylisothiazole 1D was prepared from
5-amino-3-methylisothiazole hydrochloride and di-tert-butyl
dicarbamate in CH2Cl2 and pyridine. 3-(Boc-amino)-5-
methylisothiazole 1C was prepared as described in Scheme 4.
See Honma, T.; Tada, Y. Heterocycles 1977, 6, 2925±2931,
Terent'ev, A. P.; Grandreng, I. I.; Shibiryakova, D. V.; Kost,
A. N. Zhur. Obshchei. Khim. 1960, 30, 2925±2931.
Scheme 4. (a) KMnO4, benzene±acetone, rt, 67%; (b) ClCO2Et, Et3N
then NaN3, H2O±acetone; (c) t-BuOH, toluene, rt, 70%.
12. Kozikowski, A. P.; Adamczyk, M. J. Org. Chem. 1983, 48,
366±372.
13. Yiannios, C. N.; Karabinos, J. V. J. Org. Chem. 1963, 28,
3246±3248.