Potent and orally bioavailable non-peptide antagonists at the human bradykinin B1 receptor based on a 2-alkylamino-5-sulfamoylbenzamide core

Article abstract of DOI:10.1021/jm049747g

The bradykinin B₁ receptor is rapidly induced after inflammation or tissue trauma and appears to play an important role in the maintenance of hyperalgesia in inflammatory conditions. Here, we describe the optimization process to identify novel,

Full text of DOI:10.1021/jm049747g

4642
J . Med. Chem. 2004, 47, 4642-4644
Letters
Ch a r t 1. Bradykinin B1 Antagonist Lead 1
P oten t a n d Or a lly Bioa va ila ble
Non -P ep tid e An ta gon ists a t th e Hu m a n
Br a d yk in in B1 Recep tor Ba sed on a
2-Alk yla m in o-5-su lfa m oylben za m id e Cor e
Timothy J . Ritchie,* Edward K. Dziadulewicz,
Andrew J . Culshaw, Werner Mu¨ller,†
Gillian M. Burgess,‡ Graham C. Bloomfield,§
Gillian S. Drake, Andrew R. Dunstan,§
David Beattie,§ Glyn A. Hughes, Pam Ganju,
Peter McIntyre, Stuart J . Bevan, Clare Davis, and
Mohammed Yaqoob
While BK and KD have high affinity for constitutive
BK B2 receptors, these kinins have low affinity for BK
B1 receptors, which are generally present at low levels
under normal conditions. However, B1 receptors are
rapidly induced and expressed in peripheral tissues and
cells within 2-5 h after tissue injury or inflammation
and are activated by the metabolic fragments des-Arg9-
BK and des-Arg10-KD.2
Novartis Institute for Medical Sciences, 5 Gower Place,
London WC1E 6BS, U.K.
Received March 31, 2004
Compelling evidence suggests that the BK B1 receptor
plays an important role in chronic nociceptive pain
associated with inflammatory conditions.3 In animal
models of persistent nociceptive pain, it has also been
demonstrated that peptide B1 receptor antagonists are
effective in reversing inflammatory hyperalgesia.4 Thus,
a potent, selective, orally bioavailable B1 antagonist
should have significant potential as a novel therapy for
inflammatory pain.
Abstr a ct: The bradykinin B1 receptor is rapidly induced after
inflammation or tissue trauma and appears to play an
important role in the maintenance of hyperalgesia in inflam-
matory conditions. Here, we describe the optimization process
to identify novel, potent non-peptide human B1 receptor
antagonists based on a 2-alkylamino-5-sulfamoylbenzamide
core. Optimized derivatives are selective, functional B1 an-
tagonists with low nanomolar affinity and exhibit oral bio-
availability in animals.
Several classes of non-peptide antagonists for BK B2
receptors have been disclosed in the scientific litera-
ture,5,6 and more recently the discovery of non-peptide
antagonists for the B1 receptor have been described.7,8
In this Letter, we describe our own efforts in the
optimization of a micromolar lead compound to afford
potent, selective, and orally bioavailable non-peptide BK
B1 receptor antagonists based on the 2-alkylamino-5-
sulfamoylbenzamide core.
An in-house collection of non-peptide bradykinin B2
antagonists, generated in our previous research that
afforded bradyzide and related analogues,9-11 was as-
sayed for binding affinity at the cloned human brady-
kinin B1 receptor expressed in HEK293 cells. This
screening process identified the 4-alkylamino-3-nitroben-
zene sulfonamide 1 (Chart 1) as an interesting lead,
having single-digit micromolar binding affinity (ex-
pressed as the inhibition constant KI ( SEM) at both
human B1 and B2 receptors, expressed in HEK293 and
Cos-7 cells, respectively. 1 also behaved as a functional
antagonist at human B1 receptors expressed in Cos-7
cells, inhibiting the Ca2+ efflux induced by des-Arg10-
KD with IC50 ) 6.2 µM.
Chronic pain affects a significant proportion of people
worldwide and causes unnecessary suffering, disability,
and a poor quality of life. There is a great medical need
for new medicines to treat chronic nociceptive pain
primarily because of the side effects that are frequently
observed with existing treatments such as nonsteroidal
antiinflammatory drugs (NSAIDs) and opioids.
The kinins are potent 9-10 amino acid peptide
hormones involved in inflammatory, vascular, and pain
processes and are released following inflammation,
tissue damage, or other noxious stimulation.1 In hu-
mans, the two endogenous kinins bradykinin (BK, Arg-
Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg) and kallidin (KD,
Lys-Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg) are formed
by proteolytic cleavage of high- and low-molecular-
weight kininogen precursor proteins by plasma or tissue
kallikrein (serine protease) enzymes, respectively.
The kinin peptides exert their biological effects by
activating two distinct BK receptors (termed B1 and B2),
which belong to the rhodopsin superfamily of G-protein-
coupled receptors (GPCRs) but have low (36%) sequence
homology to one another. Across species, there are also
considerable differences in the amino acid sequences of
the BK B1 receptors.
To rapidly investigate the importance of the bis-amine
and the benzhydryl moieties in 1, analogues were
prepared. Thus, in separate reactions, eight different
polar amines (see Chart 2) were coupled with com-
mercially available 4-chloro-3-nitrobenzenesulfonyl chlo-
ride in THF in the presence of Et3N to form the
corresponding sulfonamide intermediates, which were
each used crude for the subsequent nucleophilic aryl
chloride displacement (DMSO at 120 °C) with nine
* To whom correspondence should be addressed. Phone: +44 20
7333 2168. Fax: +44 20 7387 4116. E-mail: tim.ritchie@pharma.
novartis.com.
† Present address: Novartis Pharma AG, Basel, Switzerland.
‡ Present address: Pfizer Central Research, Sandwich, CT13 9NJ ,
U.K.
§ Present address: Novartis Horsham Research Centre, Wimble-
hurst Road, Horsham RH12 5AB, U.K.
10.1021/jm049747g CCC: $27.50 © 2004 American Chemical Society
Published on Web 08/10/2004

Letters
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 19 4643
Ch a r t 2. Amines Used for Initial Array of Analogues of 1
Sch em e 1. Synthesis of Piperidine-4-carboxylic Acid
Analogue 4a
a
Conditions: (a) (COCl)2, PhMe, DMF, room temp; (b) 3,3′-
iminobis(N,N′-dimethylaminopropylamine), CH2Cl2; (c) H2, 10%
Pd/C, MeOH; (d) 4-chloro-3-nitrobenzenesulfonyl chloride, NEt3,
THF; (e) 2,2′-diphenylethylamine, DMSO, 150 °C.
Ta ble 1. Binding Affinity of Nitro Group Replacements
Ch a r t 3. More Potent Analogues of 1 from First Array
compd
R
B1 KI (µM)
5
6
7
8
NO2
Cl
C(O)NMe2
0.144 ( 0.022
>1
0.136 ( 0.022
0.012 ( 0.003
Ch a r t 4. Amino Acid Linkers Used in Second Array
C(O)morpholine
eridine-2-carboxylic acid, and the aromatic amino acids
as spacers was not useful, incorporation of the 3-
aminopropionic acid, piperidine-3-carboxylic acid, and
piperidine-4-carboxylic acid spacers increased receptor
binding affinity relative to the simple ethylene spacer.
In particular, the piperidine-4-carboxylic acid derivative
of 2 (4, Scheme 1) exhibited KI ) 0.018 µM.
In parallel to the optimization of the sulfonamide-
bis-amine spacer group, analogues were prepared in
which the undesirable aromatic nitro group was re-
placed with alternative substituents. In this investiga-
tion, the 3-aminopropionic acid spacer was used with
the 2,2-diphenylethyl group (Table 1). While the chloro
derivative 6 had low affinity for B1 receptors, the
tertiary amide 7 exhibited a binding affinity similar to
that of the parent nitro analogue 5. Although primary
and secondary amide derivatives were found to be
inactive (as was the parent carboxylic acid), the utility
of a tertiary amide as a nitro replacement was confirmed
when morpholine was used in place of dimethylamine.
This change resulted in 8, which had an affinity similar
to that of 4.
separate hydrophobic amines (Chart 2). The 71 new
derivatives obtained by this process were screened for
binding affinity at the human B1 receptor and compared
with that of 1.
The results indicated that while the majority of the
members of the array were equiactive with 1 or less
potent (particularly when the bis-amine motif was
absent), a 10-fold increase in binding affinity was
observed in two analogues where (i) an ethylene spacer
had been introduced between the sulfonamide and the
bis-amine and (ii) the benzhydryl group had been
replaced by the 2,2-diphenylethyl or the dibenzylmethyl
group (2 and 3, Chart 3). It is interesting that neither
of these modifications, in the absence of the other,
increased B1 receptor binding affinity over that observed
with 1.
Subsequently a hybrid molecule was prepared, which
combined the optimal amino acid spacer (piperidine-4-
carboxylic acid used in 4) and nitro replacement (mor-
pholine amide used in 8). This analogue (9, Chart 5)
was found to be a highly potent B1 ligand, with a binding
KI of 0.001 µM. 9 also behaved as a functional antago-
nist in Cos-7 cells, with an IC50 of 0.012 µM.
As discussed above, analogues prepared during the
initial optimization of 1 that lacked the bis-amine motif
tended to lose binding affinity at the B1 receptor. With
the discovery of 9, we decided to reexplore the bis-amine
A subsequent array of analogues was prepared where
the ethylene spacer in 2 and 3 was replaced with a
variety of acyclic and cyclic amino acids (shown in Chart
4). For this array, 2,2-diphenylethylamine, dibenzyl-
methylamine, and 3,3-diphenylpropylamine were used
as the hydrophobic amine partners. A representative
synthesis for this array is shown in Scheme 1.
Results from the B1 binding assay indicated that
although the inclusion of methylaminoacetic acid, pip-

4644 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 19
Letters
Ch a r t 5
peptide antagonists for the human bradykinin B1 recep-
tor. The optimized derivatives are potent and selective
antagonists with reasonable oral bioavailability in
animals and as such have potential as a novel therapy
for pain associated with inflammatory conditions.
Ack n ow led gm en t. The authors thank Devnandan
Chatterjee, Wai Tsang, Lee Edwards, Fre´de´ric Ratel,
and Lucile Fisher for invaluable technical support,
Christian Guenat for obtaining the HRMS data, and
Olivier Kretz and Guy Taccard for conducting the
dog pharmacokinetics study. The authors also thank
Christopher Snell, Michael Brown, Peter Kipfer, J ulian
Arbuckle, Reg Docherty, Steven Phagoo, Ximena Nun˜ez,
Wai Lee, Andrew Davis, Michael Webb, and Elsa
Phillips.
Sch em e 2a
Su p p or tin g In for m a tion Ava ila ble: 1H NMR spectra
(400 MHz), HPLC and HRMS characterization data for all
compounds, elemental analyses for 1, 11, and 12, and details
of human B1 receptor binding and functional bioassays. This
material is available free of charge via the Internet at http://
pubs.acs.org.
a
Refer en ces
Conditions: (a) (COCl)2, PhMe, DMF, room temp; (b) mor-
pholine, NEt3, THF; (c) Zn, AcOH; (d) NaNO2, HCl, AcOH; (e) SO2,
CuCl2, AcOH, H2O; (f) isonipecotic acid, Na2CO3, dioxane-water;
(g) 2,2-diphenylethylamine, DMSO, 120 °C; (h) isopropyl chloro-
formate, NMM, THF, -40 °C, 40 min, followed by amine.
(1) Regoli, D.; Barabe´, J . Pharmacology of Bradykinin and Related
Kinins. Pharmacol. Rev. 1980, 32, 1-46.
(2) Marceau, F.; Hess, F.; Bacharov, D. R. The B1 Receptor for
Kinins. Pharmacol. Rev. 1998, 50, 357-85.
(3) Couture, R.; Harrison, M.; Vianna, R. M.; Cloutier, F. Kinin
Receptors in Pain and Inflammation. Eur. J . Pharmacol. 2001,
429, 161-176.
region to find a smaller monobasic amine to replace the
iminobis(N,N′-dimethylpropylamine).
(4) Belichard, P.; Landry, M.; Faye, P.; Bachvarov, D. R.; Bouthillier,
J .; Pruneau, D.; Marceau, F. Inflammatory hyperalgesia induced
by zymosan in the plantar tissue of the rat: effect of kinin
receptor antagonists Immunopharmacology 2000, 46, 139-147.
(5) Heitsch, H. Non-Peptide Antagonists and Agonists of the Brady-
kinin B2 Receptor. Curr. Med. Chem. 2002, 9, 913-928.
(6) Gurrath, M. Peptide-Binding G Protein-Coupled Receptors: New
Opportunities for Drug Design. Curr. Med. Chem. 2001, 8, 1605-
1648.
To expedite this, the advanced intermediate 10 was
prepared according to the synthetic route shown in
Scheme 2. The carboxyl group of 10 was then activated
via the corresponding mixed anhydride and coupled
with a range of cyclic and acyclic amines.
It soon became apparent that it was possible to
replace the bis-amine with a number of cyclic and acyclic
monoamines to afford analogues that retained a re-
spectable level of B1 binding affinity. As anticipated, the
oral absorption profiles of some of the monoamines were
superior to those of 9. For example, while 9 could not
be detected in plasma after oral administration to rats,
the two piperazine derivatives 11 and 12 (Chart 5) were
found to exhibit reasonable oral absorption. This im-
provement was thought to be due to the reduction in
overall molecular weight and the lower basicity of the
piperazine nitrogens (pKa of 6.5 and 7.6 for 11 and 12,
respectively). Thus, after an oral dose of 7 mg/kg,
administered as a suspension in 0.5% methyl cellulose,
the hydrochloride salts of 11 and 12 exhibited Cmax of
913 and 196 nM and oral bioavailabilities of 42% and
15%, respectively. After intravenous administration to
rats (0.7 mg/kg), the half-lives for 11 and 12 were
estimated at 161 and 37 min, respectively. In dogs, a
20 mg/kg dose of 11 exhibited an oral bioavailability of
35% (Cmax ) 10.3 µM) when administered as an aqueous
hydroxypropyl â-cyclodextrine inclusion complex.
11 and 12 also exhibited at least 500-fold selectivity
for human bradykinin B1 receptors over a panel of 30
G-protein-coupled receptors (including human BK B2
receptors), 7 ion channels (including sodium, potassium,
and calcium channels), and cyclooxygenase enzymes.
In conclusion, we have described the optimization of
a micromolar lead 1 to afford a novel series of non-
(7) Bock, M. G.; Hess, J . F.; Pettibone, D. J . Bradykinin-1 receptor
antagonists. Annu. Rep. Med. Chem. 2003, 38, 111-120.
(8) Gougat, J .; Ferrari, B.; Sarran, L.; Planchenault, C.; Poncelet,
M.; Maruani, J .; Alonso, R.; Cudennec, A.; Croci, T.; Guagnini,
F.; Urban-Szabo, K.; Martinolle, J .-P.; Soubrie, P.; Finance, O.;
Le Fur, G. SSR240612 [(2R)-2-[((3R)-3-(1,3-benzodioxol-5-yl)-
3-{[(6-methoxy-2-naphthyl)sulfonyl]amino}-propanoyl)amino]-3-
(4-{[2R,6S)-2,6-dimethyl-piperidinyl]methyl}phenyl)-N-isopropyl-
N-methylpropanamide hydrochloride], a new nonpeptide an-
tagonist of the bradykinin B1 receptor: Biochemical and
pharmacological characterization. J . Pharmacol. Exp. Ther.
2004, 309 (2), 661-669.
(9) Dziadulewicz, E. K.; Ritchie, T. J .; Hallett, A.; Snell, C. R.; Ko,
S. Y.; Wrigglesworth, R.; Hughes, G. A.; Dunstan, A. R.;
Bloomfield, G. C.; Drake, G. S.; Brown, M. C.; Lee, W.; Burgess,
G. M.; Davis, C.; Yaqoob, M.; Perkins, M. N.; Campbell, E. A.;
Davis, A. J .; Rang, H. P. 1-(2-Nitrophenyl)thiosemicarbazides:
A Novel Class of Potent, Orally Active Non-Peptide Antagonist
for the Bradykinin B2 Receptor. J . Med. Chem. 2000, 43, 769-
771.
(10) Burgess, G. M.; Perkins, M. N.; Rang, H. P.; Campbell, E. A.;
Brown, M. C.; McIntyre, P.; Urban, L.; Dziadulewicz, E. K.;
Ritchie, T. J .; Hallett, A.; Snell, C. R.; Wrigglesworth, R.; Lee,
W.; Davis, C.; Phagoo, S. B.; Davis, A. J .; Phillips, E.; Drake, G.
S.; Hughes, G. A.; Dunstan, A.; Bloomfield, G. C. Bradyzide, a
Potent Non-Peptide B2 Bradykinin Receptor Antagonist with
Long-Lasting Oral Activity in Animal Models of Inflammatory
Hyperalgesia. Br. J . Pharmacol. 2000, 129, 77-86.
(11) Dziadulewicz, E. K.; Ritchie, T. J .; Hallett, A.; Snell, C. R.;
Davies, J . W.; Wrigglesworth, R.; Dunstan, A. R.; Bloomfield,
G. B.; Drake, G. S.; McIntyre, P.; Brown, M. C.; Burgess, G. M.;
Lee, W.; Davis, C.; Yaqoob, M.; Phagoo, S. B.; Phillips, E.;
Perkins, M. N.; Campbell, E. A.; Davis, A. J .; Rang, H. P.
Nonpeptide Bradykinin B2 Receptor Antagonists: Conversion
of Rodent-Selective Bradyzide Analogues into Potent, Orally-
Active Human Bradykinin B2 Receptor Antagonists. J . Med.
Chem. 2002, 45, 2160-2172.
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