Design and synthesis of novel sulfonamide-containing bradykinin hB 2 receptor antagonists. 2. Synthesis and structure-activity relationships of α,α-cycloalkylglycine sulfonamides

550

J. Med. Chem. 2007, 50, 550-565

Design and Synthesis of Novel Sulfonamide-Containing Bradykinin hB₂Receptor Antagonists.

2. Synthesis and Structure-Activity Relationships of r,r-Cycloalkylglycine Sulfonamides

Daniela Fattori,*^,†Cristina Rossi,^†Christopher I. Fincham,^†Valerio Caciagli,^†Fernando Catrambone,^†Piero D’Andrea,^†

Patrizia Felicetti,^†Martina Gensini,^‡Elena Marastoni,^†Rossano Nannicini,^‡Marielle Paris,^†Rosa Terracciano,^†

Alessandro Bressan,^†Sandro Giuliani,^‡Carlo A. Maggi,^‡Stefania Meini,^‡Claudio Valenti,^‡and Laura Quartara^‡

Menarini Ricerche, Via Tito Speri 10, 00040 Pomezia (Rome), Italy, and Menarini Ricerche, Via Rismondo 12A, 50131 Florence, Italy

ReceiVed October 3, 2006

Recently we reported on the design and synthesis of a novel class of selective nonpeptide bradykinin (BK)

B₂receptor antagonists (J. Med. Chem. 2006, 3602-3613). This work led to the discovery of MEN 15442,

an antagonist with subnanomolar affinity for the human B2 receptor (hB2R), which also displayed significant

and prolonged activity in vivo (for up to 210 min) against BK-induced bronchoconstriction in the guinea-

pig at a dose of 300 nmol/kg (it), while demonstrating only a slight effect on BK-induced hypotension.

Here we describe the further optimization of this series of compounds aimed at maximizing the effect on

bronchoconstriction and minimizing the effect on hypotension, with a view to developing topically delivered

drugs for airway diseases. The work led to the discovery of MEN 16132, a compound which, after intratracheal

or aerosol administration, inhibited, in a dose-dependent manner, BK-induced bronchoconstricton in the

airways, while showing minimal systemic activity. This compound was selected as a preclinical candidate

for the topical treatment of airway diseases involving kinin B2 receptor stimulation.

Introduction

dose, MEN 15422 had still a noticeable effect on BK-induced

hypotension (Σ% 79), indicating that the further improvement

is needed. This current paper describes our efforts to optimize

the pharmacodynamic properties of our lead compound (lower

dose, greater airway selectivity).

Bradykinin is a nonapeptide (BK: Arg¹-Pro²-Pro³-Gly⁴-Phe⁵-

Ser⁶-Pro⁷-Phe⁸-Arg⁹) generated in plasma and tissue by activa-

tion of kininogens and exerts its effects through the constitu-

tively expressed B₂receptor or through the induced B₁receptor.

In humans and guinea pigs, the kinin B₂receptors are

expressed in the upper and lower airways.^1,2In humans, BK

induces potent bronchoconstriction and cough when inhaled by

asthmatic patients³and rhinitis-like symptoms when instilled

into the nose.⁴Furthermore, BK is generated in human nasal

secretions during rhinovirus infections⁵and allergic rhinitis.⁶

On the basis of these findings, a potential therapeutic role for

kinin B₂receptor antagonists has been hypothesized in the

treatment of airway inflammatory pathologies, such as chronic

bronchial asthma⁷or perennial and seasonal allergic rhinitis.⁸

In addition, a B₂receptor is expressed also in the cardiovascular

system, where kinins exert protective effects. Several studies

have provided evidence suggesting that BK contributes to the

antihypertensive and cardioprotective effects of angiotensin-

converting enzyme inhibitors⁹and angiotensin II antagonists.¹⁰

On the basis of these considerations, it would be ideal to block

B₂receptors in the airways without affecting them at the

cardiovascular level.

We argued that the development of new, selective, human

B₂receptor (hB₂R) antagonists for topical administration would

be the best way of achieving the goal of airway selectivity. With

this aim in mind, we undertook work on the sulfonamide moiety

of Anatibant, the most advanced hB₂R antagonist, which led to

the discovery of MEN 15422 (Chart 1).¹¹In an in vivo assay in

the guinea pig, the latter, at a dose of 300 nmol/kg, was able to

lower the residual bronchoconstriction activity (Σ%)¹²to 39

following a challenge with BK (10 nmol/kg iv); at the same

dose Anatibant had a Σ% of 73 (Figure 1). However, at this

As a general working strategy, all compounds showing a

pK_ig 9.0 in a hB₂R binding assay were assessed for their ability

to antagonize a BK-induced functional response, that is, the

accumulation of inositolphosphate (IP) as an index of the

activation of the hB₂R expressed in CHO cells (hpA₂). Promising

candidates were subsequently screened in a bioassay for their

ability to antagonize the BK-induced contraction of the guinea

pig ileum longitudinal smooth muscle (GPI) to avoid any

possible problems arising from species related selectivity when

performing in vivo tests in the guinea pig. Compounds having

a pA₂(IP) g 8.7 and a pA₂(GPI) g 9 were then tested in vivo,

via intratracheal administration, for their effect on BK-induced

bronchoconstriction and hypotension.

Chemistry

The compounds described in this study are shown in Tables

1-3, and the synthetic methods for their preparation are outlined

in Schemes 1-7. Chlorosulfonic acid 1¹¹was subjected to

radical bromination under standard conditions (NBS, AIBN) to

obtain benzylbromide 2. Formation of the sulfonamides with

the corresponding R,R-aminoacids was performed in two ways,

either via a classical base-catalyzed reaction with the amino

acid methyl ester or by the addition of the amino acid pretreated

with BSA (N,O-bis-trimethylsilylacetamide). The resulting

^aAbbreviations: AcOH, acetic acid; AcCN, acetonitrile; BSA, N,O-bis-

trimethylsilylacetamide; DIAD, diisopropyl azodicarboxylate; DIPEA, di-

isopropylethyl amine; DCM, dichloromethane; DMF, dimethylformamide;

DMSO, dimethylsulfoxide; EDAC, N-(3-dimethylaminopropyl)-N′-ethyl-

carbodiimide; EtOAc, ethyl acetate; Et₂O, diethyl ether; EtOH, ethanol;

HOAt, 7-aza-1-hydroxybenzotriazole; HOBt, 1-hydroxybenzotriazole; MeOH,

methanol; NBS, N-bromosuccinimide; PyBOP, (benzotriazol-1-yloxy)-

tripyrrolidinophosphonium hexafluorophosphate; TFA, trifluoroacetic acid;

THF, tetrahydrofuran; TMSCl, trimethylsilyl chloride.

* To whom correspondence should be addressed. Phone: +39 06

91184522. Fax: +39 06 9106137. E-mail: dfattori@menarini-ricerche.it.

^†Menarini Ricerche, Pomezia (Rome), Italy.

^‡Menarini Ricerche, Florence, Italy.

Published on Web 01/06/2007

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 551

Figure 1. In vivo activity of compounds 41-46 and reference compounds after intratracheal administration at 300 nmol/kg. For details, see the

Experimental Section. b ) Σ% (BC), bronchoconstriction, see ref 12. c ) Σ% (BP), blood pressure, see ref 12.

Chart 1. Competitive, Nonpeptidic hB₂Receptor Antagonists Anatibant and MEN 15442

methyl esters (3a, 3b, and 3c) or free acids (3d, 3e, and 3f)

were reacted with 2,4-dimethyl-quinolin-8-ol in the presence

of a base and, in the case of methyl esters, then subjected to

basic hydrolysis to obtain acids 4a-f (Scheme 1). These were

coupled with either functionalized piperazines (19, 21, 26, 38,

and 40; Scheme 2, route a) or with a monoprotected piperazine

residue (Scheme 2, route b). In the latter case, removal of the

protecting group (Boc or Fmoc) gave intermediates 6, which

were coupled with the chosen acids. The carboxylic acids with

suitably protected basic and/or charged groups were derived

from L-ornithine (Scheme 3), L-â-lysine (Scheme 4), L-lysine

(Scheme 5), or simple ω-amino acids of differing chain lengths

(Scheme 6). The functionalized piperazines were mainly two

kinds: dimethylamines (9, 18, 19, 24, and 31) generated via

reductive amination with aq HCHO in the presence of H₂/Pd

or NaCNBH₃or trimethylammonium salts (10, 12, 14, 21, 26,

29, 32, 34, and 40) obtained via treatment of the amines with

O-methyl-N,N′-diisopropylurea or dimethylsulfate. Finally, a

guanidino moiety was introduced into compound 36 using 1,3-

di-Boc-2-(trifluoromethylsulfonyl)guanidine (Goodman’s re-

agent) and alkylated via a Mitsunobu reaction (Scheme 7). In

all cases, removal of the protecting groups and HPLC purifica-

tion gave the final products.

15422 with the slightly more bulky R,R-cyclopentaneglycine

residue. Comparison of the in vitro activity of these compounds

(41-46) revealed that the cyclopentaneglycine derivatives,

generally, had similar, or slightly lower, binding affinities but

slightly higher potencies in the functional assay on the hB₂

receptor (Table 1). These compounds, as well as all the others

mentioned in this paper, had a pA₂(GPI) g 9.

The in vivo activity of the compounds in the guinea pig was

measured after intratracheal administration (300 nmol/kg),

followed 5 min later by a BK challenge (10 nmol/kg iv, repeated

every 30 min for 210 min). The pulmonary insufflation pressure

and the blood pressure were monitored as a measure of BK-

induced bronchoconstriction and BK-induced hypotension,

respectively. The results are shown in Figure 1. Under these

experimental conditions, all the compounds tested were more

active than Anatibant and all showed greater activity at the

pulmonary B₂receptors compared to that at the cardiovascular

receptors, though for some compounds, for example, 45, this

too was appreciable (Σ% 51). In two out of the three cases

examined, the cyclopentaneglycine derivatives showed better

inhibition of bronchoconstriction than their dimethylglycine

counterparts (see Figure 1, 41 vs 44 and 43 vs 46).

This small improvement led us to investigate further the new

series, and several modifications were made to the substituents

on the piperazine ring (Table 2). Efforts focused on modifying

the structure of these substituents so that, following intratracheal

administration, they would help the molecule to remain “stuck”

Results and Discussion

Initially we decided to explore the effect, in a small set of

analogues, of replacing the R,R-dimethylglycine group of MEN

552 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

Table 1. Binding and In Vitro Functional Activity on the hB₂Receptor

and Functional Activity on the Guinea-Pig Ileum (GPI) B₂Receptor for

Compounds 41-46

showed very good inhibition of BK-induced bronchoconstriction

(Σ% 24), with a minimal effect on BK-induced hypotension.

Therefore, we decided to use this molecule as the starting

point for a final set of compounds in which a heteroatom was

inserted into the central cyclic R,R-disubstituted amino acid to

increase the polar surface area in this region of the molecule

(Table 3). Again, all the compounds showed outstanding binding

affinities, and the introduction of a tetrahydropyran ring (72)

significantly enhanced the pA₂in comparison to compound 63.

Those new compounds with a pA₂g 9 were tested in vivo, and

this confirmed the excellent activity of compound 72 (MEN

16132; Figure 3). The activity of this compound remained high

even at lower doses; at 30 nmol/kg it showed a Σ% of 39 against

BK-induced bronchoconstriction, and at an even lower dose

(10 nmol/kg, i.t.) it maintained a Σ% of 63, while any significant

effect on BK-induced hypotension was undetectable (Figure 4).

At all three doses the inhibitory effect lasted for at least

210 min.

All the compounds reported in this paper were also tested

for their affinity at the hB1 receptor and all showed a pK_ivalue

of <5. Finally, given the presence of a trimethylammonium

moiety in MEN 16132 (72), the compound was tested, at 1 µM,

for any activity on the muscarinic or nicotinic receptors; none

was found.

Conclusions

In this paper we have described the extensive work carried

out on MEN 15422, the lead compound from our first paper.¹¹

Changes to the amino acid used to form the central sulfonamide

unit and to the nature and distribution of the polar groups in

the terminal chain produced a compound, MEN 16132 (72),

which was 100× more active in vivo. This new molecule, when

administered locally, was a potent and long lasting antagonist

of BK-induced bronchoconstriction in the airways, devoid of

any significant effects on BK-induced hypotension.¹⁶It could

be potentially useful in the treatment of chronic airway diseases

involving the proinflammatory activity of BK and is under study.

Additional pharmacological results will be reported in due

course.

^apK_ifor inhibition of specific binding of [³H]-BK to hB₂receptor in

stably transfected CHO cells membrane preparations. ^bpA₂for the hB₂-

mediated accumulation of inositol monophosphate in stably transfected

CHOdhfr-/hB₂R cells. ^cpA₂for the antagonism of BK-induced contractile

responses in guinea-pig ileum longitudinal smooth muscle. For details see

the Experimental Section.

to the surface of the respiratory tract,¹⁴with a minimal amount

being released into circulation to interact with the cardiovascular

receptors. A similar strategy has been used for the muscarinic

receptor antagonists ipratropium, flutropium, and tiotropium.¹⁵

Thus, it was decided to increase the polarity of the molecule

via the introduction of positive charges through the use of very

basic functional groups (protonated at physiological pH) and

quaternary ammonium salts. These groups have the added

advantage of rendering the molecules poorly bioavailable thus

avoiding absorption of the compounds following accidental

ingestion during intratracheal or aerosol administration.

Carboxylic acids containing amines, ammonium salts, and

guanidines at varying distances and geometries were linked to

the piperazine moiety (Table 2). All the new compounds, with

the single exception of compound 50, showed subnanomolar

binding affinities, and all had good activity in the functional

test (pA₂g 8.2). Best of all was ammonium salt 63, with a pK_i

of 10.3 and a pA₂of 9.3. The combination of a primary amine

and a quaternary ammonium salt at the correct distance from

one another appeared to be the key for both high binding affinity

and high antagonist potency (compound 63 vs compounds 49

and 61). The majority of the compounds were then screened in

vivo, at a dose of 100 nmol/kg, to assess their potency and

selectivity of action in inhibiting bronchoconstriction over

hypotension (Figure 2a). At this dose, only compound 63

Experimental Section

(A) Chemistry. Commercial chemicals and solvents were of

reagent grade and used without further purification.

Merck silica gel (Kieselgel 60) was used for analytical thin-

layer chromatography (TLC, F254 plates) and flash chromatography

(230-400 mesh).

Purity was evaluated by analytical HPLC using a 600 E Waters

pump coupled to a Jasco 875 UV detector and a Merck-Hitachi

D-2500 integrator, or a system comprising a Jasco PU-980 pump,

LG-980-02 gradient unit, a UV-975 UV/vis detector, and a Merck-

Hitachi D-2500 integrator, a Beckman System Gold apparatus, or

an Agilent 1100 analytic HPLC system. The solvents were (A) water

+ 0.1% TFA and (B) AcCN + 0.1% TFA, flow 1 mL/min.

System A: Column Vydac RP-18, 5 µm, 250 × 4.6 mm, λ )

220 nm; from 95% to 20% solvent (A) in 25 min. System B:

Column Symmetry 300 RP-18, 250 × 4.6 mm, λ ) 220 or 254

nm; from 80% to 20% solvent (A) in 20 min. System C: Column

Agilent Zorbax Eclipse XDB C8, 5 µm, 150 × 4.6 mm; λ ) 210,

240 nm; from 80% to 20% solvent (A) in 20 min. System D:

column Jupiter RP-18, 5 µm, 150 × 4.6 mm, λ ) 210, 240 nm;

from 80% to 20% solvent (A) in 20 min.

Preparative reverse phase HPLC was performed on a Waters

600E apparatus with a Jasco 874 UV detector or on a Waters Delta-

Prep 3000 apparatus. The mobile phases were the same as for the

analytical systems. Gradient elution was employed. The columns

used were a SymmetryPrep C18, 7 µm, 19 × 300 mm, a Hibar

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 553

Table 2. Binding and In Vitro Functional Activity on the hB₂Receptor of Compounds 47-67

^aSee Table 1. ^bSee Table 1. For details see the Experimental Section.

Lichrosorb RP-18, 7 µm, 25 × 250 mm, a Vydac C18, 10 µm,

22 × 250 mm or Jupiter, 15 µm, 250 × 21.2 mm. Peak detection

was at 220 and 254 nm. Chemical yields are not optimized.

(50 × 3 mm, 3 µm) or a ThermoFinnigan LCQ equipped with APCI

or ESI source.

1-(3-Bromomethyl-2,4-dichloro-benzenesulfonylamino)-cyclo-

pentanecarboxylic Acid Methyl Ester (3a). A mixture of 2 (150

mg, 0.445 mmol), 1-aminocyclopentancarboxylic acid methyl ester

(87 mg, 0.490 mmol), and K₂CO₃(150 mg, 0.980 mmol) in DMF

(15 mL) was stirred at room temperature for 18 h. At the end of

the reaction (TLC control) the solution was diluted with water (150

mL) and extracted with EtOAc (3 × 50 mL). The organic layer

was washed with 1 N HCl (3 × 50 mL), water (50 mL), and brine

(50 mL), dried over Na₂SO₄, and concentrated to afford a solid

(100 mg) as a mixture of 3a and the corresponding benzyl chloride

NMR experiments were recorded on a Varian Gemini 200 mod.

J200 HC, a Varian 300 MHz spectrometer (equipped with a 5 mm

inverse probe), a Bruker Avance 400 MHz, or a Bruker Avance

600 MHz machine and are referenced to residual solvent signals:

CDCl₃(δ 7.26) or DMSO-d₆(δ 2.49). Chemical shifts are reported

in δ units (parts per million) and are assigned as singlets (s),

doublets (d), doublets of doublets (dd), triplets (t), quartet (q),

quintet (quin), multiplets (m), broad signals (br), or very broad

signals (v br). Coupling constants (J) are reported in hertz (Hz).

1

in a ratio of 3:7. H NMR (300 MHz, CDCl₃): δ 8.06-7.97 (1H,

Mass spectra were recorded using a Waters Alliance 2795 HPLC

system fitted with a UV-PDS 996 diode array detector, a ZMD

mass spectrometer, and a GL Science Inertsil ODS-3 column

m), 7.54-7.48 (1H, m), 5.44 (1H, s), 4.94 (1.4H, s, CH₂Cl), 4.82

(0.6H, s, CH₂Br), 3.66-3.60 (3H, m), 2.52-2.03 (2H, m), 2.01-

554 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

Scheme 2^a

Table 3. Binding and In Vitro Functional Activity on the hB₂Receptor

of Compounds 68-76

^aReagents: (a) EDAC, HOAt, R′CO-piperazine; (b) HOAt or HOBt,

EDAC, Boc-piperazine (5a-5c, 5e, 5f) or Fmoc-piperazine (5d); (c) HCl/

dioxane; (d) piperidine, DMF; (e) R′CO₂H, EDAC, HOBt, DIPEA.

Scheme 3^a

^aSee Table 1. ^bSee Table 1. For details see the Experimental Section.

Scheme 1^a

^aReagents: (a) HCHO, H₂, 10% Pd/C, MeOH; (b) O-methyl-N,N′-

diisopropylisourea, MeOH; (c) Me₂SO₄, NaOH.

mL) at 5 °C. Stirring was continued at this temperature for 19.5 h,

after which time HPLC analysis showed almost complete consump-

tion of the sulfonyl chloride. The AcCN was removed in vacuo,

the residue was diluted with water (80 mL), and the mixture was

filtered under reduced pressure. The solid was washed with water

(3×), air-dried for 30 min, then transferred to a tared flask and

dried under high vacuum to constant weight to give 3b as an off-

white solid in a 3:1 mixture with the corresponding benzyl chloride

(2.981 g, 75% yield). HPLC purity: system B, t_R17.46 min

(12.4%), 17.88 min (83.8%). ¹H NMR (300 MHz, CDCl₃): δ 7.99-

7.91 (1H, m), 7.53-7.46 (1H, m), 5.30 (1H, br s), 4.94 (0.5H, s,

CH₂Cl), 4.81 (1.5H, s, CH₂Br), 3.71-3.49 (7H, m), 2.25-2.07 (2H,

m), 1.99-1.81 (2H, m). MS m/z calcd for C₁₄H₁₆BrCl₂NO₅S,

458.93; found, 460.1, 462.1, 464.1 [M + H]⁺. MS m/z calcd for

C₁₄H₁₆Cl₃NO₅S, 414.98; found, 438.3, 440.1 [M + Na]⁺. HPLC

purity: system B, t_R17.46 min (12.4%), 17.88 min (83.8%).

^aReagents: (a) NBS, AIBN, CCl₄; (b) amino acid methyl ester, base;

(c) amino acid, BSA, TMSCl, THF; (d) 2,4-dimethyl-quinolin-8-ol, KI, base;

(e) LiOH, THF/H₂O or NaOH, THF/H₂O; (f) HCl/dioxane; (g) HCHO,

BH₃-CN resin, MeOH; (h) Ac₂O, DIPEA, DMF.

4-(3-Bromomethyl-2,4-dichloro-benzenesulfonylamino)-tet-

rahydro-thiopyran-4-carboxylic Acid Methyl Ester (3c). A

solution of 1-amino-tetrahydrothiopyran carboxylic acid methyl

ester hydrochloride (100 mg, 0.47 mmol) in 5% NaHCO₃(0.7 mL)

was added to a solution of sulfonyl chloride 2 (132 mg, 0.39 mmol)

in AcCN (2 mL). The resulting mixture was stirred at room

temperature. At the end of the reaction (HPLC control), the solvents

were distilled off in vacuo, and the residue was dissolved in EtOAc,

washed with 1 M KHSO₄(3×), dried over Na₂SO₄, and concen-

trated to obtain 3c (88 mg) together with the corresponding benzyl

chloride in the ratio 7:3 (50% yield). MS m/z calcd for C₁₄H₁₆-

1.86 (2H, m), 1.80-1.51 (4H, m). MS m/z calcd for C₁₄H₁₆BrCl₂-

NO₄S, 442.9; found, 460.8, 462.9, 464.8 [M + NH₄]⁺. MS m/z

calcd for C₁₄H₁₆Cl₃NO₄S, 399.0; found, 416.9, 419.0, 421.0 [M +

NH₄]⁺. HPLC purity: system A, t_R17.24 min (68.4%), 17.62 min

(27.0%).

4-(3-Bromomethyl-2,4-dichloro-benzenesulfonylamino)-tet-

rahydro-pyran-4-carboxylic Acid Methyl Ester (3b). A solution

of 5% NaHCO₃(53 mL, 31.5 mmol) was added at a rate of 10

mL/h to a solution of the sulfonyl chloride 2 (2.91 g, 8.61 mmol)

and the 4-amino-tetrahydropyran-4-carboxylic acid methyl ester

hydrochloride (2.059 g, 10.52 mmol) in AcCN/water 10:1 (58.3

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 555

Scheme 4^a

^aReagents: (a) H₂, 10% Pd/C, MeOH; (b) Goodman’s reagent; (c) HCHO, NaCNBH₃; (d) Fmoc-piperazine, EDAC, HOBt, DIPEA; (e) H₂, 10% Pd/C,

AcOH; (f) MeI; (g) piperidine/DMF; (h) HCl/dioxane.

Scheme 5^a

^aReagents: (a) H₂, 10% Pd/C, MeOH; (b) Me₂SO₄, NaOH; (c) TFA, CH₂Cl₂, then HCHO, NaCNBH₃.

Scheme 6^a

EtOAc, washed with water (4×), and dried over Na₂SO₄to obtain

crude 3d (86 mg), which was used as such in the following reaction.

MS m/z calcd for C₁₈H₂₃BrCl₂N₂O₆S, 543.98; found, 542.8, 544.8,

546.8 [M - H⁺]^-. HPLC purity: system B (100-0% A in 30 min),

t_R21.00 min (20.0%), 21.26 min (80.0%).

1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-ben-

zenesulfonylamino]-cyclopentanecarboxylic Acid (4a). A solution

of 2,4-dimethyl-quinolin-8-ol (42 mg, 0.24 mmol), 3a (and the

corresponding benzyl chloride in the ratio 3:7; 100 mg, 0.24 mmol),

KI (43 mg, 0.26 mmol), and K₂CO₃(74 mg, 0.48 mmol) in dry

acetone (10 mL) was refluxed for 7 h under nitrogen. At the end

of the reaction (HPLC control), the mixture was poured over acetate

buffer at pH 4.2 (100 mL) and extracted with EtOAc (3 × 50 mL).

The organic layers were washed with water (3 × 70 mL) and brine,

dried over Na₂SO₄, filtered, and concentrated to afford 1-[2,4-

dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-benzenesulfonyl-

amino]-cyclopentanecarboxylic acid methyl ester (100 mg). A

portion of this material (50 mg, 0.093 mmol) was dissolved in THF/

MeOH/water (3:2:1; 6 mL) together with LiOH (24 mg, 0.1 mmol),

and the resulting mixture was refluxed for 4 h. At the end of the

reaction (HPLC control), the reaction was cooled to room temper-

ature, the organic solvents were removed in vacuo, and the residue

was diluted with water (10 mL). The pH was made acidic (ca. 5)

by the cautious addition of 1 N HCl, and the solution was extracted

with EtOAc (3 × 25 mL). The organic extracts were combined,

washed with water (3 × 30 mL) and brine, dried over Na₂SO₄,

^aReagents: (a) TFA, CH₂Cl₂; (b) Me₂SO₄, NaOH.

BrCl₂NO₄S₂, 474.9; found, 492.9, 495.0, 496.9 [M + NH₄]⁺. HPLC

purity: system B t_R16.21 min (16.3%), 16.54 min (77.4%).

4-(3-(Bromomethyl)-2,4-dichloro-phenylsulfonamido)-1-(tert-

butoxycarbonyl)piperidine-4-carboxylic Acid (3d). A suspension

of 4-Boc-1,1-aminopiperidinylcarboxylic acid (195 mg, 0.80 mmol)

in dry THF (2.4 mL) was stirred at reflux temperature under

nitrogen, and BSA (0.59 mL, 2.4 mmol) and TMSCl (0.31 mL,

2.4 mmol) were added. Refluxing was continued for an additional

2 h until dissolution of the amino acid was complete. Then a

solution of sulfonyl chloride 2 (134 mg, 0.40 mmol) in dry THF

(3 mL) was added dropwise with stirring. At the end of the reaction

(HPLC control, RP-C18), the mixture was diluted with MeOH/

H₂O (1:1) and concentrated in vacuo to afford an oily yellow

residue. This was diluted with EtOAc, washed with water (2×)

and 1 M NaHSO₄(3×), dried over Na₂SO₄, and concentrated in

vacuo to afford an oil that solidified upon drying. This solid

consisted of a mixture of 3d and the corresponding benzyl chloride

in a 4:1 mixture (83 mg, 41% yield). It was dissolved in acetone/

DMF (1:2; 1.6 mL), NaBr (130 mg, 1.27 mmol) was added, and it

was stirred at room temperature to convert the benzyl chloride into

the benzyl bromide. At the end of the reaction (HPLC control),

the solvent was distilled off in vacuo, the residue was dissolved in

1

filtered, and concentrated to afford crude 4a (41 mg). H NMR

(300 MHz, CDCl₃): δ 8.13 (1H, d, J ) 8.6 Hz), 7.68-7.50 (3H,

m), 7.25 (1H, d, J ) 7.8 Hz), 7.21 (1H, s), 5.80 (1H, s), 5.60 (2H,

556 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

Figure 2. In vivo activity of compounds 49-67 after intratracheal administration at 100 nmol/kg. For details, see the Experimental Section; 66 and

67 were tested at 300 nmol/kg. b ) Σ% (BC), bronchoconstriction, see ref 12. c ) Σ% (BP), blood pressure, see ref 12.

Scheme 7^a

^aReagents: (a) Goodman’s reagent; (b) DIAD, PPh₃, 3-dimethylaminopropanol; (c) HCl/dioxane; (d) MeI.

s), 2.71 (3H, s), 2.58 (3H, s), 2.24-1.57 (8H, m). HPLC purity:

system A, 80.0%, t_R10.07 min.

(1H, dd, J ) 8.3, 8.3 Hz), 7.37 (1H, d, J ) 8.3 Hz), 7.30 (1H, s),

5.58 (2H, s), 3.50-3.38 (2H, m), 3.35-3.10 (2H, m), 2.64 (3H,

s), 2.56 (3H, s), 1.90-1.75 (4H, m). MS m/z calcd for C₂₄H₂₄-

Cl₂N₂O₆S, 538.1; found, 539.2, 541.2 [M + H]⁺. HPLC purity:

system B, 95.0%, t_R11.63 min.

4-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-ben-

zenesulfonylamino]-tetrahydro-pyran-4-carboxylic Acid (4b).

2,4-Dimethyl-quinolin-8-ol (2.253 g, 13.02 mmol) was dissolved

in a 1.5 M solution of Bu₄NOH (8.6 mL, 13 mmol) within 30 min.

The resulting intense orange solution was added to Bu₄NBr (347

mg, 1.08 mmol) and compound 3b (as a 4:1 mixture with the

corresponding benzyl chloride; 4.88 g, 10.8 mmol) in AcCN (20

mL). The mixture immediately turned green, and after about 4 h,

a white solid separated from the solution. The course of the reaction

was monitored by HPLC. After 24 h, the mixture was diluted with

water (50 mL) and filtered, and the resulting solid was washed

with water (100 mL), petroleum ether (15 mL), and MeOH (15

mL) and then dried in vacuo to yield the desired compound in the

form of methyl ester (4.68 g). A batch of the methyl ester so

obtained (5.46 g, 9.86 mmol) was suspended in AcCN (50 mL),

and the mixture was heated to reflux. A 15% aq NaOH solution

(52 mL, 197 mmol) was added, and complete solution occurred to

give a biphasic system, which was refluxed for 24 h. At the end of

the reaction (HPLC control), the mixture was cooled to room

temperature and concentrated in vacuo to remove the AcCN, and

the resulting cloudy solution was carefully acidified with 4 N HCl

to pH 4-5. After stirring for 1 h, a fine white precipitate formed,

which was filtered off, washed with water (100 mL) and Et₂O (30

4-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-ben-

zenesulfonylamino]-tetrahydro-thiopyran-4-carboxylic Acid (4c).

A mixture of 2,4-dimethyl-quinolin-8-ol (35 mg, 0.20 mmol) and

NaH (6.0 mg, 0.24 mmol) in DMF (3 mL) was stirred at room

temperature for 45 min. Then a solution of 3c (and the correspond-

ing benzyl chloride in ratio 7:3; 91 mg, 0.20 mmol) and KI

(60 mg, 0.40 mmol) in DMF (3 mL) was added dropwise. At the

end of the reaction (HPLC control), the solvent was distilled off in

vacuo, and the residue was dissolved in EtOAc. The organic solution

was washed with pH 4.2 buffer (3×), dried over Na₂SO₄, and

concentrated to give an orange solid (88 mg, 78% yield). This was

dissolved in dioxane (3.0 mL) and heated to 100 °C, and a 5% aq

NaOH solution (2.9 mL) was added while stirring. At the end of

the reaction (HPLC control), the solution was concentrated in vacuo,

and the orange residue was partitioned between EtOAc and pH

4.2 buffer. The two phases were separated, and the organic one

was dried over Na₂SO₄and concentrated to obtain 4c (61 mg, 72%

yield) as a white solid. MS m/z calcd for C₂₄H₂₄Cl₂N₂O₅S₂, 554.05;

found, 555.0, 557.0 [M + H]⁺. HPLC purity: system B, 85%; t_R

10.43 min.

1

mL), and dried in vacuo to afford 4b in 97% yield (5.45 g). H

4-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-ben-

zenesulfonylamino]-piperidine-1,4-dicarboxylic Acid Mono-tert-

butyl Ester (4d). A mixture of 1,1-dimethyl-quinolin-8-ol (28 mg,

NMR (400 MHz, DMSO-d₆): δ 8.64 (1H, br s), 8.03 (1H, d, J )

8.8 Hz), 7.75 (1H, d, J ) 8.8 Hz), 7.67 (1H, d, J ) 8.3 Hz), 7.46

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 557

Figure 3. In vivo activity of compounds 68, 70, and 72 after intratracheal administration at 100 nmol/kg. For details, see the Experimental Section.

b ) Σ% (BC), bronchoconstriction, see ref 12. c ) Σ% (BP), blood pressure, see ref 12.

Figure 4. In vivo activity of compound 72 after intratracheal administration on bronchoconstriction and hypotension induced by repeated challenges

with BK (10 nmol/kg iv) in anaesthetized guinea pigs.¹⁶

0.16 mmol) and LiOH (10 mg, 0.4 mmol) in dry AcCN (2.5 mL)

was stirred at room temperature for 1.5 h under nitrogen. Then a

solution of crude 3d (86 mg, 0.16 mmol) in AcCN/DMF (2:1)

(2 mL) was added dropwise, and the resulting mixture was stirred

at room temperature. At the end of the reaction (HPLC control),

MeOH/water (1:1; 3 mL) was added, stirring was continued for an

additional 15 min, and then the solvents were distilled off in vacuo.

The residue was partitioned between EtOAc and pH 4.2 buffer.

The two phases were separated, and the organic one was washed

again with buffer (5×). The organic layer was dried over Na₂SO₄,

filtered, and concentrated in vacuo. The residue was triturated with

Et₂O to afford crude 4d (70 mg, 70% yield). MS m/z calcd for

C₂₉H₃₃Cl₂N₃O₇S, 637.14; found, 638.1, 640.0 [M + H]⁺. HPLC

purity: system B, (100-0% A in 30 min), 70%, t_R17.59 min.

2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-N-[1-

(piperazine-1-carbonyl)-cyclopentyl]-benzenesulfonamide (6a).

EDAC (981 mg, 5.12 mmol) and HOBt (629 mg, 5.12 mmol) were

added to a solution of acid 4a (2.43 g, 4.66 mmol) in CH₂Cl₂

(50 mL) stirred in an ice bath. The resulting mixture was stirred

for an additional hour, then N-Boc-piperazine (697 mg, 5.12 mmol)

and DIPEA (0.88 mL, 5.12 mmol) were added, and stirring was

continued at room temperature overnight. Then the mixture was

diluted with CH₂Cl₂(20 mL) and washed with 0.1 N KHSO₄

(50 mL), 0.1 N NaOH (50 mL), and brine (50 mL), dried over

Na₂SO₄, filtered, and concentrated in vacuo. The crude product was

purified by flash chromatography to give the Boc-protected

derivative in 99% purity. This was suspended in EtOAc (15 mL),

and 4 N HCl in dioxane (20 mL) was added dropwise while stirring

at room temperature. At the end of the reaction (HPLC control),

the mixture was concentrated in vacuo, dissolved in water, and

lyophilized to give 6a (2.15 g, 73% yield) as a white solid in the

1

form of the hydrochloride salt. H NMR (300 MHz, CDCl₃): δ

8.94 (2H, s), 8.7 (s, 1H), 8.34 (1H, br s), 8.04-8.01 (1H, d), 7.83-

7.81 (1H, d), 7.61-7.45 (4H, m), 5.56 (2H, s), 3.16 (4H, br s),

2.75 (3H, s), 2.69 (3H, s), 2.65 (3H, s), 1.93 (2H, m), 1.73 (2H,

m), 1.42 (4 H, m). MS m/z calcd for C₂₈H₃₂Cl₂N₄O₄S, 591.56;

found, 592.2 [M + H]⁺. HPLC purity: system B (100-0% A in

30 min), 94%, t_R10.37 min.

2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-N-[4-

(piperazine-1-carbonyl)-tetrahydro-pyran-4-yl]-benzenesulfona-

mide (6b). EDAC (51 mg, 0.27 mmol) was added to a chilled

solution of acid 4b (105 mg, 0.195 mmol) and HOAt (37 mg, 0.27

mmol) in DCM (6.0 mL). The resulting mixture was stirred for 30

min at 0 °C and then N-Boc-piperazine (66 mg, 0.35 mmol) was

added. Stirring was continued at 0 °C for 30 min then at room

temperature. At the end of the reaction (HPLC control), the DCM

558 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

was removed in vacuo, and the residue was partitioned between

EtOAc (50 mL) and pH 4.2 buffer (50 mL). The layers were

separated, and the organic phase was washed with buffer (50 mL).

The aqueous washes were combined and re-extracted with EtOAc

(50 mL), then the organic extracts were combined and washed with

5%NaHCO₃(2 × 50 mL) and brine (2 × 50 mL), dried over Na₂-

SO₄, and filtered, and the filtrate was concentrated in vacuo to give

as the hydrochloride salt (176 mg). A portion of this product (50

mg, 0.08 mmol) was dissolved in MeOH (4.5 mL), and 37% aq

HCHO (30.7 µL, 0.41 mmol) was added, together with NaCNBH₃

polystyrene resin (38 mg, 4.3 mmol/gr, 0.16 mmol), and stirring

was continued at room temperature. At the end of the reaction

(HPLC control), the resin was filtered off and the residual solution

was concentrated in vacuo, with the help of toluene (4×), to obtain

crude 4e.

1

crude 5b. H NMR (300 MHz, CDCl₃): δ 8.02 (1H, d, J ) 8.7

Hz), 7.64 (1H, d, J ) 8.3 Hz), 7.50 (1H, d, J ) 8.6 Hz), 7.39 (1H,

t, J ) 8.0 Hz), 7.20 (1H, d, J ) 7.6 Hz), 7.15 (1H, s), 5.73 (2H,

s), 5.52 (1H, s), 3.87-3.69 (4H, m), 3.63-3.43 (8H, m), 2.69 (3H,

s), 2.64 (3H, s), 2.21-2.10 (2H, m), 1.65-1.52 (2H, m), 1.47 (9H,

s). MS m/z calcd for C₃₃H₄₀Cl₂N₄O₇S₂, 706.2; found, 707.2 [M +

H]⁺. HPLC purity: system B, 95.9%, t_R15.22 min. HCl/dioxane

(4 N, 4 mL) was added at room temperature to a solution of 5b

(138 mg, 0.195 mmol) in DCM (4 mL). At the end of the reaction

(HPLC control), the solvents were removed in vacuo, and the

residue was triturated with Et₂O, filtered, washed with Et₂O, and

dried under a stream of N₂to give 6b (131 mg, 98%) as a pale

Crude 4e (40 mg, 0.07 mmol) in dry DMF (1 mL) was stirred at

room temperature. HOAt (98 mg, 0.07 mmol), DCC (14.8 mg, 0.07

mmol), and Boc-piperazine (13 mg, 0.07 mmol) were added in that

order and stirring was continued overnight. Na₂CO₃(5%, 5 mL)

was added and stirring was continued for an additional 30 min.

The mixture was then extracted with EtOAc (3×) and the organic

layer was washed with 5% Na₂CO₃(2×). The EtOAc layer was

acidified with diluted HCl to pH 4, washed with water, dried over

Na₂SO₄, and concentrated under reduced pressure. The resulting

amorphous solid was dissolved in THF, cooled for 3 days at -20

°C, filtered to eliminate the residual DCU, and concentrated again

to obtain crude 5e. This material was dissolved in dry EtOAc

(1 mL), 4 N HCl in dioxane (5 mL) was added, and the resulting

mixture was stirred for 30 min. The solvents were distilled off and

the residue was dried in vacuo to obtain crude 6e (114 mg), which

was used as such without further purification.

1

yellow solid. H NMR (300 MHz, DMSO-d₆): δ 9.73-9.50 (2H,

m), 8.92 (1H, s), 8.46-8.12 (4H, m), 7.34-7.08 (2H, m), 5.68

(2H, br s), 4.15-3.77 (4H, m), 3.27-2.97 (6H, m), 2.83 (6H, br

s), 1.98-1.77 (2H, m), 1.74-1.54 (2H, m). MS m/z calcd for

C₂₉H₃₄Cl₂N₄O₄S, 606.15; found, 607.2, 609.2 [M + H]⁺. HPLC

purity: system B, 94.9%, t_R9.46 min.

N-[1-Acetyl-4-(piperazine-1-carbonyl)-piperidin-4-yl]-2,4-

dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-benzenesulfona-

mide (6f). A portion of 4-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-piperidine-4-carboxylic acid

hydrochloride salt (see preparation for 6e; 60 mg, 0.098 mmol)

was dissolved in dry DMF (1 mL). Acetic anhydride (9.3 µL, 0.098

mmol) and DIPEA (50 µL, 0.29 mmol) were added while stirring

at room temperature. After 4 h (HPLC control), water (2 mL) was

added and the solvents were distilled off in vacuo. Toluene was

added, and the mixture was concentrated again (4×). The solid

residue was triturated with Et₂O to obtain crude 4f (78 mg). This

crude product (0.098 mmol) was dissolved in dry DMF (2 mL)

and then EDAC (20.7 mg, 0.108 mmol), HOAt (14.7 mg, 0.108

mmol), DIPEA (32 µL, 0.098 mmol), and Boc-piperazine (20.1

mg, 0.108 mmol) were added in that order while stirring at room

temperature. After 12 h (HPLC control), 5% NaHCO₃(5 mL) was

added and stirring was continued for an additional 30 min. Then

EtOAc was added and the two phases were separated. The organic

phase was washed with 5% NaHCO₃(3×) and pH 4.2 buffer (3×),

dried over Na₂SO₄, and concentrated. Flash chromatographic

purification (silica, CHCl₃, then CHCl₃/MeOH 99:1, then CHCl₃/

MeOH 98:2) afforded pure 5f (19 mg, 0.025 mmol). This was

dissolved in dry dioxane (1 mL), and 4 N HCl in dioxane (2 mL)

was added while cooling in an ice bath. At the end of the reaction

(HPLC control), the solvents were distilled off and the residue was

triturated with Et₂O to obtain 6f (18 mg, quantitative). MS m/z calcd

for C₃₀H₃₅Cl₂N₅O₅S, 647,17; found, 648.1, 650.1 [M + H]⁺.

(S)-2-tert-Butoxycarbonylamino-5-dimethylamino-pentano-

ic Acid (9). BocOrn-OH (8) was suspended in MeOH (100 mL),

and 37% aq HCHO (1.6 mL, 0.02 mmol) was added, followed by

10% Pd/C (200 mg). The reaction mixture was stirred under H₂

for 3 h, then the catalyst was filtered off, and the filtrate was

concentrated to dryness. Et₂O (100 mL) was added to the resulting

oil, and stirring was continued to obtain a white solid, which was

filtered off and washed with Et₂O. A second precipitation was

performed by dissolving the solid in the minimum amount of MeOH

(3 mL) and then adding Et₂O (100 mL). The desired product was

finally collected by filtration, following a further wash with Et₂O,

as a highly hygroscopic white solid that was stored under nitrogen.

Total yield, 1.60 g (60%). ¹H NMR (400 MHz, DMSO-d₆): δ 6.49

(1H, d), 6.05 (1H, d), 3.75-3.65 (1H, m), 2.70-2.53 (2H, m), 2.39

(6H, s), 1.70-1.41 (4H, m), 1.38 (9H, s). ¹³C NMR (100 MHz,

DMSO-d₆): δ 175.27, 155.94, 78.54, 58.21, 54.74, 43.99, 30.23,

29.09, 22.5. MS m/z calcd for C₁₂H₂₄N₂O₄, 260.0; found, 261.0

[M + H]⁺.

2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-N-[4-

(piperazine-1-carbonyl)-tetrahydro-thiopyran-4-yl]-benzene-

sulfonamide (6c). A solution of 4c (45 mg, 0.083 mmol) and

DIPEA (28 µL, 0.083 mmol) in DMF (2 mL) was stirred at room

temperature. PyBOP (43 mg, 0.083 mmol) was added, followed

by N-Boc-piperazine (15 mg, 0.083 mmol). At the end of the

reaction (HPLC control), the solvent was distilled off in vacuo,

and the orange residue was dissolved in EtOAc and washed with

acetate buffer, pH 4.2 (3×). The organic layer was dried over Na₂-

SO₄, filtered, and concentrated to obtain crude 5c (45 mg). MS

m/z calcd for C₃₃H₄₀Cl₂N₄O₆S₂, 722.18; found, 723.1, 725.1 [M +

H]⁺. This crude product was dissolved in TFA/CH₂Cl₂(1:1; 2 mL)

and stirred at room temperature. After 1 h, Et₂O was added with

stirring and a white solid precipitated. This was filtered off and

dried under nitrogen to give crude 6c as trifluoroacetate salt (26

mg), which was used as such without further purification. MS m/z

calcd for C₂₈H₃₂Cl₂N₄O₄S₂, 622.12; found, 623.1, 625.1 [M + H]⁺.

4-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-ben-

zenesulfonylamino]-4-(piperazine-1-carbonyl)-piperidine-1-car-

boxylic Acid tert-Butyl Ester (6d). A solution of 4d (70 mg, 0.11

mmol) in DMF (3 mL) was stirred at room temperature. EDAC

(25 mg, 0.13 mmol) and HOBt (18 mg, 0.13 mmol) were added.

Fmoc-piperazine hydrochloride (38 mg, 0.110 mmol) was dissolved

in DMF (1 mL), DIPEA was added (0.04 mL, 0.26 mmol), and

the resulting solution was added dropwise to the solution of the

active ester. The resulting mixture was stirred overnight. The

solvents were distilled off in vacuo, and the residue was partitioned

between EtOAc and pH 4.2 buffer. The organic layer was washed

with buffer (2×) and then dried over Na₂SO₄, concentrated, and

triturated with Et₂O to obtain crude 5d as a white solid (76 mg,

75% yield). MS m/z calcd for C₄₈H₅₁Cl₂N₅O₈S, 927.28; found,

928.2, 930.2 [M + H]⁺. A solution of 5d (76 mg, 0.081 mmol) in

piperidine/DMF 1:10 (3 mL) was stirred at room temperature for

1 h. The solution was concentrated in vacuo, and the resulting

yellow oil was triturated with Et₂O to obtain 6d as a white solid

(47 mg, 81% yield). MS m/z calcd for C₃₃H₄₁Cl₂N₅O₆S, 705.22;

found, 706.2, 708.2 [M + H]⁺.

2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-N-[1-

methyl-4-(piperazine-1-carbonyl)-piperidin-4-yl]-benzenesulfona-

mide (6e). A solution of 4d (173 mg, 0.27 mmol) in dry dioxane

(1 mL) was cooled to 0 °C, 4 N HCl in dioxane (10 mL) was added,

and the mixture was warmed to room temperature. At the end of

the reaction (2 h, HPLC control), the solvents were distilled off

under reduced pressure and the residue was triturated with Et₂O

(3×) to obtain crude 4-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-

yloxymethyl)-benzenesulfonylamino]-piperidine-4-carboxylic acid

(S)-2-tert-Butoxycarbonylamino-5-trimethylammonium-pen-

tanoic Acid (10). Amino acid 9 (1.16 g, 4.37 mmol) was dissolved

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 559

in MeOH (5 mL). O-Methyl-N,N′-diisopropylisourea (0.87 mL, 4.81

mmol) was added, and the reaction mixture was stirred at room

temprature for 18 h (MS-HPLC control). Solvent removal under

reduced pressure, followed by addition of water (30 mL), caused

the precipitation of the diisopropylurea, which was filtered off. The

aqueous layer was washed with EtOAc (3 × 30 mL) to remove

the unreacted O-methyl-N,N′-diisopropylisourea, and then the water

was again distilled off under reduced pressure. Absolute EtOH

(3 × 50 mL) was added and then removed under reduced pressure

until a white foam formed. Et₂O (50 mL) was added, and the

resulting suspension was stirred at room temperature for 1 h.

Ammonium salt 10 was finally collected, after filtration and washing

with Et₂O, as a highly hygroscopic white solid that was stored under

of solid CO₂to the water bath. At the end of the deprotection

reaction (HPLC control), the solution was degassed with nitrogen

for 20 min, while maintaining the temperature at 0 °C, and then

filtered. The filtrate was concentrated in vacuo to give the free amine

as the acetate salt. This crude product was purified by flash

chromatography (CHCl₃/MeOH/AcOH 85:10:5) to obtain the pure

amine as the acetate salt. MS m/z calcd for C₃₀H₄₀N₄O₅, 536.30;

found, 537.3 [M + H]⁺. HPLC purity: system B, >99%, t_R16.42

min.

This salt was dissolved in CH₂Cl₂/AcOH 9:1 (10 mL), and 37%

aq HCHO (220 µL, 2.71 mmol) was added. The resulting solution

was agitated (150 oscillations/min) at room temperature for 20 min,

(polystyrylmethyl)trimethylammonium cyanoborohydride (339 mg,

1.39 mmol) was added, and the agitation was continued at room

temperature. At the end of the reaction (HPLC and TLC control),

the resin was filtered off under nitrogen and washed with CH₂Cl₂

(3 × 10 mL), and the filtrates were combined and concentrated in

vacuo to give 19 (318 mg) as an off-white solid after concentration

from toluene (3×). A portion of this product was used in the next

step without further purification. MS m/z calcd for C₃₂H₄₄N₄O₅,

564.33; found, 565.3 [M + H]⁺.

{(S)-4-tert-Butoxycarbonylamino-6-[4-(9H-fluoren-9-ylmethox-

ycarbonyl)-piperazin-1-yl]-6-oxo-hexyl}-trimethyl-ammonium

trifluoroacetate (20). Methyl iodide (160 µL, 2.57 mmol) was

added to a solution of 19 (260 mg, 0.419 mmol) in CH₂Cl₂/MeOH

(2:1; 6 mL) and stirring at room temperature was continued for 2

days. At the end of the reaction (HPLC control), the solvents were

removed in vacuo and the residue was triturated with Et₂O. The

resulting off-white solid was filtered under nitrogen, washed with

Et₂O (3 × 6 mL), and dried under a stream of nitrogen to give

crude 20. This crude product was purified by preparative HPLC to

obtain pure 20 (118 mg, 40%) as the trifluoroacetate salt. ¹H NMR

(300 MHz, CDCl₃): δ 7.78-7.74 (2H, m), 7.60-7.53 (2H, m),

7.42-7.37 (2H, m), 7.37-7.28 (2H, m), 5.88-5.71 (1H, br s), 4.49

(2H, d, J ) 6.4 Hz), 4.29-4.20 (1H, t, J ) 6.4 Hz), 4.02-3.86

(1H, br s), 3.64-3.29 (10H, m), 3.16 (9H, m), 2.70-2.47 (2H, m),

1.99-1.69 (3H, m), 1.68-1.49 (1H, m), 1.41 (9H, s). MS m/z calcd

for C₃₃H₄₇N₄O₅, 579.35; found, 579.2 [M + H]⁺. HPLC purity:

system B, 98.6%, t_R16.90 min.

[(S)-4-tert-Butoxycarbonylamino-6-oxo-6-piperazin-1-yl-hexyl]-

trimethyl-ammonium Trifluoroacetate (21). A solution of 20 (84

mg, 0.119 mmol) in 20% piperidine/DMF (2.4 mL) was stirred at

room temperature. At the end of the deprotection reaction (HPLC

control), the solvents were removed in vacuo and the residue was

concentrated twice from DMF under reduced pressure. The resulting

white solid was triturated with Et₂O and the supernatant was

removed by pipet. The residue was dissolved in MeOH and

concentrated again to obtain 21 in the form of the trifluoroacetate

salt (59 mg), which was used as such without any further

purification.

1

nitrogen (1.10 g, 90% yield). H NMR (400 MHz, DMSO-d₆): δ

6.49 (1H, d), 3.45 (1H, m), 3.29 (2H, m), 3.03 (9H, s), 1.75-1.58

(4H, m), 1.38 (9H, s). ¹³C NMR (100 MHz, DMSO-d₆): δ 172.38,

155.73, 78.19, 66.3, 55.15, 52.6, 29.9, 28.06, 18.7. MS m/z calcd

for C₁₃H₂₇N₂O₄, 275.0; found, 275.0 [M]⁺.

(S)-(4-tert-Butoxycarbonylamino-1-carboxy-butyl)-trimethyl-

ammonium (12). Amino acid 11 (0.100 g, 0.40 mmol) was

dissolved in 10% NaOH (4 mL), Me₂SO₄(0.4 mL) was added,

and the reaction mixture was stirred for 30 min at 0 °C and then

for 30 min at room temperature (MS-HPLC control). The solution

was neutralized with 0.5 N HCl, the solvent was removed under

reduced pressure, and the residue was triturated with Et₂O to obtain

12, which was used without further purification. MS m/z calcd for

C₁₃H₂₇N₂O₄, 275.37; found, 275.4 [M]⁺.

(S)-(1-Carboxy-butyl)-1,4-ditrimethyl-ammonium (14). Amino

acid 13 monochloride (0.200 g, 1.20 mmol) was dissolved in 10%

NaOH (5 mL), Me₂SO₄(0.5 mL) was added, and the reaction

mixture was stirred for 30 min at 0 °C and for 30 min at room

temperature (MS-HPLC control). The solution was neutralized with

0.5 N HCl, the solvent was removed under reduced pressure, and

the residue was triturated with Et₂O to obtain 14, which was used

without further purification. MS m/z calcd for C₁₁H₂₆N₂O₂, 218.34;

found, 218.5 [M]⁺.

(S)-6-Amino-3-tert-butoxycarbonylamino-hexanoic Acid (16).

A solution of Boc-â-Lys(Z)-OH (210 mg, 0.55 mmol) in MeOH

(20 mL) was stirred under H₂in the presence of 10% Pd/C (50

mg). At the end of the reaction (TLC control, silica, CHCl₃/MeOH

9:1), the solution was filtered through Celite and the filtrate was

concentrated in vacuo to obtain 16 (130 mg, 96%) as a colorless

oil. MS m/z calcd for C₁₁H₂₆N₂O₂, 218.34; found, 219.2 [M + H]⁺.

(S)-6-[(tert-Butoxycarbonimidoylimino-tert-butoxycarbony-

lamino-methyl)-amino]-3-tert-butoxycarbonylamino Hexanoic

Acid (17). A suspension of 16 (130 mg, 0.52 mmol) in CH₂Cl₂

(5 mL) was treated with BSA (0.40 mL, 1.56 mmol) until complete

solution occurred. Goodman’s reagent (600 mg, 1.56 mmol) was

added, and stirring was continued at room temperature for 24 h.

At the end of the reaction (TLC control, silica, CHCl₃/MeOH 9:1),

the mixture was concentrated under reduced pressure, and the crude

residue was purified by flash chromatography (silica, CH₂Cl₂/MeOH

9:1) to obtain 17 (112 mg, 44%). MS m/z calcd for C₂₂H₄₀N₄O₈,

488.594; found, 489.6 [M + H]⁺.

(S)-3-tert-Butoxycarbonylamino-6-dimethylamino-hexanoic

Acid (18). A solution of 16 (113 mg, 0.45 mmol) in dioxane (10

mL) was treated with 37% aq HCHO (250 µL, 3.0 mmol), and

NaCNBH₃(75 mg, 1.2 mmol) in water (3 mL) was added. At the

end of the reaction (TLC control, silica, AcCN/water 5:15), the

mixture was concentrated under reduced pressure, and the crude

product was purified by flash chromatography (silica, CH₃CN/H₂O

1:4) to obtain 18 (65 mg, 55%). MS m/z calcd for C₁₃₁H₂₆N₂O₄,

274.36; found, 275.4 [M + H]⁺.

4-[(S)-6-Benzyloxycarbonylamino-3-tert-butoxycarbonylamino-

hexanoyl]-piperazine-1-carboxylic Acid 9H-Fluoren-9-ylmethyl

Ester (22). Commercially available Boc-â-Lys(Z)-OH DCHA (987

mg, 1.76 mmol) was partitioned between EtOAc (100 mL) and 1

N HCl (100 mL). The layers were separated, and the organic phase

was washed with water (2 × 50 mL) and brine (2 × 50 mL), dried

(Na₂SO₄), and concentrated under reduced pressure to give 15 as

the free acid. A solution of acid 15 (447 mg, 1.17 mmol), HOBt

(217 mg, 1.42 mmol), and EDAC (286 mg, 1.49 mmol) in CH₂Cl₂

(4 mL) was stirred at 0 °C for 30 min. Then Fmoc-piperazine

hydrochloride (412 mg, 1.19 mmol) was added, followed by DIPEA

(420 µL, 2.41 mmol), and stirring was continued at 0 °C for 30

min and then at room temperature. At the end of the reaction (HPLC

control), the solvent was distilled off in vacuo, and the residue was

partitioned between EtOAc (50 mL) and 1 N HCl (50 mL). The

layers were separated and the organic phase was washed with 1 N

HCl (50 mL). The combined acid washes were back-extracted with

EtOAc (50 mL), then the organic extracts were combined and

washed with 5% NaHCO₃(2 × 50 mL), water (50 mL), and brine

(50 mL), dried over Na₂SO₄, and concentrated in vacuo to afford

4-[(S)-3-tert-Butoxycarbonylamino-6-dimethylamino-hexanoyl]-

piperazine-1-carboxylic Acid 9H-Fluoren-9-ylmethyl Ester (19).

Pd/C (10%, 45 mg) was wetted with three drops of water, suspended

in AcOH (2 mL), and added, under nitrogen, to a solution of 22

(400 mg, 0.596 mmol) in AcOH (4 mL). The resulting mixture

was cooled in a water bath to 10 °C and degassed with nitrogen

for 10 min. Hydrogen was then bubbled through the mixture while

the temperature was maintained below 15 °C by periodic addition

1

crude 22 (694 mg, 88%) as a pale yellow foam. H NMR (300

560 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

MHz, DMSO-d₆): δ 7.94-7.80 (2H, m), 7.68-7.60 (2H, m), 7.46-

7.29 (9H, m), 7.26-7.19 (1H, br t, J ) 5.5 Hz), 6.70-6.63 (1H, d,

J ) 8.8 Hz), 4.99 (2H, s), 4.46-4.37 (2H, d, J ) 6.1 Hz), 4.33-

4.25 (1H, t, J ) 6.2 Hz), 3.77-3.65 (1H, m), 3.45-3.25 (8H, m),

3.02-2.91 (2H, m), 2.48-2.28 (2H, m), 1.48-1.28 (13H, m).

HPLC purity: system B, 83.6%, t_R23.26 min.

(0.67 mL) was added, followed by a solution of NaCNBH₃(164

mg, 2.6 mmol) in water. At the end of the reaction (TLC control,

RP-18, AcCN/water 1:1), the solvents were distilled off under

reduced pressure and the residue was purified by RP-flash chro-

matography to obtain 31 (127 mg, 60%). MS m/z calcd for

C₁₀H₂₂N₂O₂, 202.30; found, 203.4 [M + H]⁺.

4-[(S)-3-Amino-6-benzyloxycarbonylamino-hexanoyl]-piper-

azine-1-carboxylic Acid 9H-Fluoren-9-ylmethyl Ester (23). A 4

N solution of HCl in dioxane (4 mL, 8 mmol) was added dropwise

to a solution of 22 (232 mg, 0.346 mmol) in CH₂Cl₂(4 mL), and

stirring was continued for 40 min at room temperature. The solvents

were removed in vacuo, and the residue was dissolved in MeOH,

concentrated (2×), dissolved in toluene/MeOH (1:1), and concen-

trated again. The resulting oil was triturated with Et₂O to obtain a

solid, which was filtered off under nitrogen to give 23 (198 mg,

(R)-2,6-Bis-trimethylammonium-hexanoic Acid (32). A mix-

ture of 31 (70 mg, 0.3 mmol) in MeOH (3 mL), NaOH (70 mg,

1.75 mmol) in water (1.5 mL), and Me₂SO₄(0.5 mL, 0.45 mmol)

was stirred for 30 min in an ice bath and then for 30 min at room

temperature. It was neutralized by the dropwise addition of 1 N

HCl, and the solvents were distilled off in vacuo. The residue was

washed with petroleum ether to leave the ammonium salts 32 as a

mixture with NaCl, which was used, as such, without further

purification. MS m/z calcd for C₁₂H₃₀N₂O₂, 232.2; found, 231.4

[M + H]⁺.

General Synthesis of the ω Ammonium Salts 34 (Scheme 6).

A solution of the commercially available Boc-protected derivatives

33 (0.43 mmol) in CH₂Cl₂(10 mL) and TFA (1 mL) was stirred at

room temperature for 1 h. The solvents were distilled off in vacuo,

and the residue was dissolved in 10% NaOH (4 mL) and Me₂SO₄

(0.4 mL) was added. After 30 min, the mixture was neutralized by

the dropwise addition of 1 N HCl. The solvents were distilled off

in vacuo, and the residue was washed with petroleum ether to leave

the ammonium salts 34 as a mixture with NaCl, which were used,

as such, without further purification.

4-{tert-Butoxycarbonylamino-[(E)-tert-butoxycarbonylimino]-

methyl}-piperazine-1-carboxylic Acid tert-Butyl Ester (36). A

solution of tert-butyl 1-piperazinecarboxylate (252 mg, 1.35 mmol),

DIPEA (290 µL, 1.66 mmol), and Goodman’s reagent (446 mg,

1.14 mmol) in dry CH₂Cl₂(2 mL) was stirred at room temperature

overnight. At the end of the reaction (HPLC control), the solvent

was removed in vacuo and the residue was partitioned between

ethyl acetate (50 mL) and ice cooled 1 N HCl (50 mL). The layers

were separated and the organic phase was washed with 1 N HCl

(50 mL). The acid washes were combined and back-extracted with

EtOAc (50 mL). The organic extracts were combined and washed

with 5% NaHCO₃(2 × 50 mL) and brine (50 mL), dried over

Na₂SO₄, filtered, and concentrated in vacuo to obtain 36 (335 mg,

69%) as a colorless oil, which was used, as such, in the next

reactions. ¹H NMR (300 MHz, DMSO-d₆): δ 8.95 (1H, br s), 3.34

(8H, br s), 1.45-1.34 (27H, m). HPLC purity: system B, 93.3%,

t_R16.39 min.

4-{[tert-Butoxycarbonyl-(3-dimethylamino-propyl)-amino]-

[(Z)-tert-butoxycarbonylimino]-methyl}-piperazine-1-carboxy-

lic Acid tert-Butyl Ester (37). DIAD (272 µL, 1.38 mmol) was

added dropwise (at such a rate that the yellow coloration disap-

peared between each addition) to an ice-cold solution of 36 (198

mg, 0.462 mmol), triphenylphosphine (606 mg, 2.31 mmol), and

3-dimethylaminopropanol (163 µL, 1.38 mmol) in anhydrous THF

(3 mL). When the addition was complete, the cooling bath was

removed and the reaction was stirred at room temperature for 18

h. The solvent was removed in vacuo, and the residue was purified

by flash chromatography, eluting with chloroform-MeOH (9:1),

to give 37 (180 mg, 76%). MS m/z calcd for C₂₅H₄₇N₅O₆, 513.35;

found, 514.2 [M + H]⁺.

1

94%) as the hydrochloride salt. H NMR (300 MHz, DMSO-d₆):

δ 7.92-7.87 (2H, m), 7.82-7.71 (3H, br s), 7.65-7.60 (2H, m),

7.46-7.24 (10H, m), 5.01 (2H, s), 4.45-4.38 (2H, d, J ) 6.2 Hz),

4.31-4.25 (1H, t, J ) 6.2 Hz), 3.50-3.31 (8H, m), 3.04-2.94

(2H, m), 2.77-2.68 (1H, m), 2.60-2.52 (1H, m), 1.64-1.42 (4H,

m). HPLC purity: system B, 90.3%, t_R17.72 min.

4-[(S)-6-Benzyloxycarbonylamino-3-dimethylamino-hexanoyl]-

piperazine-1-carboxylic Acid 9H-Fluoren-9-ylmethyl Ester (24).

A solution of 23 (82 mg, 0.135 mmol) and 37% aq HCHO (66 µL,

0.81 mmol) in MeOH (2 mL) was stirred at room temperature for

20 min. (Polystyryl)trimethylammonium cyanoborohydride (124

mg, 4.1 mmol) was added and stirring was continued at room

temperature. At the end of the reaction (HPLC control), the resin

was filtered off and washed with MeOH (3×), and the filtrates

were combined and concentrated under reduced pressure to give

crude 24 (83 mg, 97%) as a colorless oil, which was used, as such,

without further purification. MS m/z calcd for C₃₅H₄₂N₄O₅, 598.32;

found, 599.1 [M + H]⁺.

((S)-4-Benzyloxycarbonylamino-1-{2-[4-(9H-fluoren-9-yl-

methoxycarbonyl)-piperazin-1-yl]-2-oxo-ethyl}-butyl)-trimethyl-

ammonium (25). A solution of crude 24 (77 mg, 0.12 mmol) and

MeI (80 µL, 1.3 mmol) in MeOH (2 mL) was stirred at room

temperature for five days. Because some starting amine still

remained, the solvents were distilled off in vacuo, and the residue

was dissolved in neat MeI (4 mL) and stirred at room temperature

in the dark for an additional three days. The MeI was removed in

vacuo, and the residue was purified by preparative HPLC to obtain

25 (54 mg, 62%), as a colorless oil, in the form of trifluoroacetate

salt. MS m/z calcd for C₃₆H₄₅N₄O₅, 613.34; found, 613.20 [M]⁺.

[(S)-4-Benzyloxycarbonylamino-1-(2-oxo-2-piperazin-1-yl-

ethyl)-butyl]-trimethyl-ammonium Iodide (26). A solution of 25

(54 mg, 0.074 mmol) in 20% piperidine/DMF (4 mL) was stirred

at room temperature for 30 min, then the solvents were removed

under reduced pressure, and the residue was concentrated from

DMF (2×). The resulting white crystalline solid was triturated with

Et₂O (4 mL × 3) to remove the benzofulvene-piperidine adduct,

and the residue was dried in vacuo to obtain 26 (39 mg, quantitative)

as a white solid, which was used as such. MS m/z calcd for

C₂₁H₃₅N₄O₃, 391.27; found, 391.3 [M]⁺.

[(R)-5-tert-Butoxycarbonylamino-1-carboxy-pentyl]-trimethyl-

ammonium (29). A solution of 27 (150 mg, 0.38 mmol) in MeOH

(10 mL) was stirred under H₂in the presence of 10% Pd/C (100

mg). When the deprotection reaction was complete, the mixture

was filtered through Celite and the solution was concentrated in

vacuo. The crude product was dissolved in 10% NaOH (4 mL),

Me₂SO₄(0.4 mL) was added, and stirring was continued in an ice

bath for 30 min and then at room temperature for an additional 30

min. The reaction was neutralized by the dropwise addition of 5 N

HCl, and the solution was concentrated in vacuo to obtain crude

29, which was used as such without any further purification. MS

m/z calcd for C₁₄H₃₀N₂O₄, 290.41; found, 290.5 [M ]⁺.

N-(3-Dimethylamino-propyl)-piperazine-1-carboxamidine (38).

A solution of 4 N HCl in dioxane (3 mL) was added to 37 (60 mg,

0.117 mmol) in MeOH (1 mL), and the reaction was stirred at room

temperature overnight. The solvents were distilled off under reduced

pressure to give 38 (40 mg, quantitative yield) as the hydrochloride

1

salt, which was used without further purification. H NMR (400

MHz, DMSO-d₆): δ 10.79 (1H, br s), 9.59 (2H, br s), 8.48 (1H, br

s), 8.12 (2H, br s), 3.80-3.40 (8H, m), 3.38-3.29 (2H, m), 3.15-

3.05 (2H, m), 2.75 (6H, s), 2.05-1.87 (2H, m).

(R)-2,6-Bis-dimethylamino-hexanoic Acid (31). A solution of

30 (300 mg, 1.09 mmol) in CH₂Cl₂(5 mL) and TFA (1 mL) was

stirred at room temperature for 2 h. At the end of the reaction (TLC

control), the solvents were distilled off in vacuo to obtain crude

31. This was dissolved in dioxane (5 mL), and 37% aq HCHO

[3-(tert-Butoxycarbonyl-{[(E)-tert-butoxycarbonylimino]-pip-

erazin-1-yl-methyl}-amino)-propyl]-trimethyl-ammonium (39).

A solution of 37 (50 mg, 0.097 mmol) and MeI (60 µL, 0.58 mmol)

in Et₂O was stirred overnight at room temperature. A white

precipitate formed, which was filtered off and dried in vacuo to

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 561

give crude 39 (50 mg, 70%), which was used, as such, in the next

reaction. MS m/z calcd for C₉H₂₁N₅, 199.30; found, 528.2 [M]⁺.

Trimethyl-{3-[(piperazine-1-carboximidoyl)-amino]-propyl}-

ammonium (40). A solution of 4 N HCl in dioxane (3 mL) was

added to crude 39 (50 mg, 0.099 mmol) in MeOH (1 mL), and the

reaction was stirred at room temperature overnight. The solvents

were removed in vacuo, and the residue was triturated with Et₂O

to give crude 40 as a highly hygroscopic solid, which was used

1.61 (4H, m), 1.53-1.25 (10H, m). MS m/z calcd for C₃₅H₄₆-

Cl₂N₈O₅S, 760.27; found, 761.1 [M + H]⁺. HPLC purity: System

B, 95%, t_R) 8.80 min.

2,4-Dichloro-N-{1-[4-((S)-2,6-diamino-hexanoyl)-piperazine-

1-carbonyl]-cyclopentyl}-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonamide Trifluoroacetate Salt (47). A solution of

Boc-D-Lys(Boc)-OH (20 mg, 0.060 mmol) in DMF (3.0 mL) was

cooled in an ice bath. EDAC (11.0 mg, 0.060 mmol) and HOAt

(8.0 mg, 0.060 mmol) were added, and the resulting mixture was

stirred for an additional hour at 0 °C. Amine 6a (30 mg, 0.040

mmol) and DIPEA (9.0 µL, 0.05 mmol) were added, and stirring

was continued at room temperature overnight. At the end of the

reaction, the solvents were distilled off in vacuo, the residue was

diluted with CH₂Cl₂(2.0 mL), and TFA (0.5 mL) added. At the

end of the Boc-deprotection reaction, solvents were distilled off in

vacuo, and the residue was purified by preparative HPLC to obtain

47 (42 mg, 96%) in the form of trifluoroacetate salt. ¹H NMR (400

MHz, DMSO-d₆): δ 8.32 (1H, s), 8.10 (3H, br s), 8.02 (1H, d),

7.80 (3H, br s), 7.77 (1H, d), 7.76-7.33 (2H, m), 7.45 (1H, t),

7.30 (2H, m), 5.68 (2H, s), 4.48 (1H, br s), 2.81 (2H, br s), 2.79

(3H, s), 2.68 (3H, s), 2.02 (2H, s), 1.80 (4H, br s), 1.45-1.60 (8H,

m). MS m/z calcd for C₃₄H₄₄Cl₂N₆O₅S, 718.25; found, 719.2 [M

+ H]⁺. HPLC purity: System B, 98%, t_R) 8.51 min.

N-{1-[4-((S)-2,6-Bis-dimethylamino-hexanoyl)-piperazine-1-

carbonyl]-cyclopentyl}-2,4-dichloro-3-(2,4-dimethyl-quinolin-8-

yloxymethyl)-benzenesulfonamide Trifluoroacetate Salt (48). A

solution of (S)-2,6-bis-dimethylaminohexanoic acid (20 mg, 0.096

mmol), EDAC (28 mg, 0.096 mmol), and HOAt (20 mg, 0.096

mmol) in DMF (5.0 mL) was cooled in an ice bath. After 1 h,

amine 6a (50 mg, 0.072 mmol) and DIPEA (11 µL, 0.072 mmol)

were added and stirring was continued overnight at room temper-

ature. At the end of the reaction (HPLC control), the solvents were

distilled off in vacuo, and the crude product was purified by

preparative HPLC to obtain 48 (50.5 mg, 63%) as a trifluoroacetate

salt. ¹H NMR (400 MHz, DMSO-d₆): δ 9.40 (1H, br s), 8.32 (1H,

s), 8.02 (1H, d), 7.77 (1H, d), 7.76-7.33 (4H, m), 5.68 (2H, s),

4.48 (1H, br s), 3.89-3.45 (8H, m), 3.18-3.04 (2H, m), 2.81 (3H,

s), 2.79 (3H, s), 2.68 (3H, s), 2.64 (3H, s), 2.09-1.28 (10H, m).

MS m/z calcd for C₃₈H₅₂Cl₂N₆O₅S, 774.31; found, 775.3 [M + H]⁺.

HPLC purity: System B, >99%, t_R) 8.65 min.

1

without further purification. H NMR (400 MHz, DMSO-d₆): δ

9.65 (2H, br s), 8.59 (1H, br s), 8.15 (2H, br s), 3.85-3.75 (4H,

m), 3.43-3.38 (4H, m), 3.36-3.28 (4H, m), 3.11 (9H, s), 2.08-

1.96 (2H, m).

N-{1-[4-((S)-2-Amino-6-dimethylamino-hexanoyl)-piperazine-

1-carbonyl]-cyclopentyl}-2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonamide Trifluoroacetate Salt (44).

A solution of the Boc-Lys(Me)₂OH (13 mg, 0.046 mmol) in DMF

(5 mL) was cooled in an ice bath. EDAC (9.0 mg, 0.046 mmol)

and HOAt (6.3 mg, 0.046 mmol) were added and stirring was

continued for 1 h. Amine 6a (25 mg, 0.031 mmol) and DIPEA (8

µL, 0.046 mmol) were added and stirring was continued at room

temperature overnight. At the end of the reaction (HPLC control),

the solvent was distilled off in vacuo, the residue was dissolved in

CH₂Cl₂(5 mL), and TFA (1 mL) was added. At the end of the

Boc-deprotection reaction (HPLC control), the solvents were

distilled off again and the crude residue was purified by preparative

1

HPLC to obtain 44 (30 mg, 88%) as the trifluoroacetate salt. H

NMR (400 MHz, DMSO-d₆): δ 9.44 (1H, br s), 8.64 (1H, s), 8.23-

8.10 (3H, br s), 8.03 (1H, d), 7.83 (1H, d), 7.75 (1H, br s), 7.69-

7.33 (8H, m), 5.59 (2H, s), 4.48 (1H, br s), 3.00 (1H, m), 2.78

(6H, s), 2.74-2.58 (4H, m), 2.60 (6H, s), 2.06-1.23 (14H, m).

MS m/z calcd for C₃₆H₄₈Cl₂N₆O₅S, 746.28; found, 747.2 [M + H]⁺.

HPLC purity: System B, 98.7%, t_R) 8.96 min.

N-{1-[4-((S)-3-Amino-6-dimethylamino-hexanoyl)-piperazine-

1-carbonyl]-cyclopentyl}-2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonamide Trifluoroacetate Salt (45).

A solution of acid 18 (9.0 mg, 0.032 mmol) in DMF (3 mL) was

cooled in an ice bath. EDAC (6.0 mg, 0.032 mmol) and HOAt

(4.0 mg, 0.032 mmol) were added, and the resulting mixture was

stirred for an additional hour at 0 °C. Then amine 6a (15 mg, 0.022

mmol) and DIPEA (5 µL, 0.032 mmol) were added and stirring

was continued for 12 h at room temperature. At the end of the

reaction (HPLC control), the solvents were distilled off in vacuo,

the residue was dissolved in CH₂Cl₂(3.0 mL), and TFA (0.7 mL)

was added. At the end of the Boc-deprotection reaction (HPLC

control), the solvents were distilled off again and the residue was

purified by preparative HPLC to obtain 45 (15 mg, 63% yield) as

[(S)-5-Amino-6-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-

piperazin-1-yl)-6-oxo-hexyl]-trimethyl-ammonium Trifluoro-

acetate Salt (49). A solution of (S)-2-Fmoc-amino-6-trimethylam-

moniumhexanoic acid (20 mg, 0.045 mmol), EDAC (13 mg, 0.067

mmol), and HOAt (9 mg, 0.067 mmol) in DMF (5.0 mL) was

cooled in an ice bath. After 1 h, amine 6a (30 mg, 0.045 mmol)

and DIPEA (7 µL, 0.045 mmol) were added and stirring was

continued overnight at room temperature. At the end of the reaction

(HPLC control), the solvents were distilled off in vacuo and the

residue was diluted with CH₂Cl₂(2.0 mL) and piperidine (1 mL).

At the end of the Fmoc-deprotection reaction, the crude product

was purified by preparative HPLC to obtain 49 (15 mg, 29%) as

1

the trifluoroacetate salt. H NMR (400 MHz, DMSO-d₆): δ 9.47

(1H, br s), 8.61 (1H, s), 8.03 (1H, d), 7.82 (1H, d), 7.81 (3H, s),

7.68 (1H, d), 7.51 (1H, t), 7.39 (1H, d), 7.32 (1H, br s), 5.55 (2H,

s), 3.52 (8H, m), 3.03-2.84 (2H, br s), 2.76 (3H, s), 2.63 (3H, s),

2.56 (3H, s), 2.03-1.34 (10H, m). MS m/z calcd for C₃₆H₄₈-

Cl₂N₆O₅S, 746.28; found, 747.3 [M + H]⁺. HPLC purity: System

B, 95%, t_R) 8.85 min.

1

the trifluoroacetate salt. H NMR (400 MHz, DMSO-d₆): δ 8.66

N-{1-[4-((S)-2-Amino-6-guanidino-hexanoyl)-piperazine-1-

carbonyl]-cyclopentyl}-2,4-dichloro-3-(2,4-dimethyl-quinolin-8-

yloxymethyl)-benzenesulfonamide Trifluoroacetate Salt (46). A

solution of Boc-L-homoarginine hydrochloride (17 mg, 0.046 mmol)

in DMF (3.0 mL) was cooled in an ice bath. EDAC (9.0 mg, 0.046

mmol) and HOAt (6.3 mg, 0.046 mmol) were added, and the

resulting mixture was stirred for an additional hour at 0 °C. Then

amine 6a (25 mg, 0.031 mmol) and DIPEA (8 µL, 0.046 mmol)

were added and stirring was continued overnight at room temper-

ature. At the end of the reaction (HPLC control), the solvents were

distilled off in vacuo, the residue was dissolved in CH₂Cl₂(3.0

mL), and TFA (1.0 mL) was added. At the end of the Boc-

deprotection reaction (HPLC control), the solvents were distilled

off again and the residue was purified by preparative HPLC to

afford 46 (27 mg, 79%). ¹H NMR (400 MHz, DMSO-d₆): δ 8.65

(1H, s), 8.14 (3H, br s), 8.04 (1H, d), 7.83 (1H, d), 7.81-7.44

(5H, m), 7.39-6.76 (3H, s), 5.50 (2H, s), 4.46 (1H, br s), 3.07

(2H, m), 2.72 (3H, s), 2.67 (3H, s), 2.03-1.91 (2H, m), 1.80-

(1H, s), 8.27-8.12 (3H, br s), 8.04 (1H, d), 7.84 (1H, d), 7.81-

7.37 (4H, m), 5.60 (2H, s), 4.60-4.42 (1H, br s), 3.70-3.42 (8H,

m), 3.24 (2H, m), 3.15 (9H, s), 2.75 (3H, s), 2.67 (3H, s), 2.04-

1.93 (2H, m), 1.82-1.22 (14H, m). MS m/z calcd for C₃₇H₅₁-

Cl₂N₆O₅S, 761.3; found, 761.4 [M]⁺. HPLC purity: System B,

98.8%, t_R) 8.61 min.

2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-N-{1-

[4-(6-guanidino-hexanoyl)-piperazine-1-carbonyl]-cyclopentyl}-

benzenesulfonamide Trifluoroacetate Salt (50). A solution of

6-Boc-amino-hexanoic acid (7.4 mg, 0.032 mmol) in DMF (2.0

mL) was cooled in an ice bath. EDAC (6.0 mg, 0.032 mmol) and

HOAt (4.3 mg, 0.032 mmol) were added, and the resulting mixture

was stirred for an additional hour at 0 °C. Amine 6a (15 mg, 0.022

mmol) and DIPEA (5.0 µL, 0.032 mmol) were added and stirring

was continued at room temperature. At the end of the reaction

(HPLC control), the solvents were distilled off in vacuo, and the

residue was treated with CH₂Cl₂(2 mL) and TFA (0.5 mL). At the

562 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

end of the Boc-deprotection reaction (HPLC control), the solvents

were distilled off in vacuo, and the residue was purified by

preparative HPLC. The resulting amine was dissolved in CH₂Cl₂,

Goodman’s reagent (15 mg, 0.037 mmol) and DIPEA (7 mg, 0.055

mmol) were added, and the resulting mixture was stirred at room

temperature overnight. At the end of the reaction (HPLC control),

the solvents were distilled off in vacuo, and the residue was purified

by preparative HPLC to obtain 50 (10 mg, 47%) as trifluoroacetate

mL) was cooled in an ice bath. After 1 h, amine 6a (50 mg, 0.11

mmol) and DIPEA (25 µL, 0.11 mmol) were added and stirring

was continued overnight at room temperature. At the end of the

reaction (HPLC control), the solvents were distilled off and the

residue was dissolved in CH₂Cl₂/TFA (3:1; 5.0 mL). After

deprotection of the Boc group, the solvents were distilled off and

the crude product was purified by preparative HPLC to afford 54

1

(21 mg, 17%) as the trifluoroacetate salt. H NMR (400 MHz,

1

salt. H NMR (400 MHz, DMSO-d₆): δ 8.44 (1H, s), 8.02 (1H,

DMSO-d₆): δ 8.35 (1H, s), 8.07 (1H, d), 8.01-7.94 (1H, m), 7.87

(2H, s), 7.80 (1H, d), 7.77-7.59 (3H, br s), 5.74 (2H, s), 4.65-

4.58 (1H, m), 3.98-3.51 (8H, m), 3.22 (9H, s), 2.91 (6H, s), 2.82-

2.80 (2H, m), 2.13-1.41 (14H, m). MS m/z calcd for C₃₇H₅₁-

Cl₂N₆O₅S, 761.3; found, 761.4 [M]⁺. HPLC purity: System B,

99.2%, t_R) 8.64 min.

d), 7.79 (1H, d), 7.76-7.32 (5H, m), 7.05-6.84 (4H, br s), 5.55

(2H, s), 3.66-3.47 (8H, m), 3.10 (2H, m), 2.68 (3H, s), 2.52 (3H,

s), 2.38-2.32 (2H, m), 2.08-1.23 (14H, m). MS m/z calcd for

C₃₅H₄₅Cl₂N₇O₅S, 745.26; found, 746.5 [M + H]⁺. HPLC purity:

System B, 96.5%, t_R) 10.17 min.

[6-(4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl}-piperazin-1-yl)-

6-oxo-hexyl]-trimethyl-ammonium Trifluoroacetate Salt (51). A

solution of (5-carboxy-pentyl)-trimethyl-ammonium (80 mg, 0.17

mmol), EDAC (32 mg, 0.17 mmol), and HOAt (23 mg, 0.17 mmol)

in DMF (5 mL) was cooled in an ice bath. After 1 h, amine 6a (30

mg, 0.04 mmol) and DIPEA (6 µL, 0.04 mmol) were added, and

stirring was continued overnight at room temperature. At the end

of the reaction (HPLC control), the solvents were distilled off in

vacuo, and the residue was purified by preparative HPLC to give

2,4-Dichloro-N-{1-[4-((S)-3,6-diamino-hexanoyl)-piperazine-

1-carbonyl]-cyclopentyl}-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonamide Trifluoroacetate Salt (55). A solution of

â-Boc-lysine (22 mg, 0.064 mmol), HOAt (87 mg, 0.043 mmol),

and EDAC (12.2 mg, 0.043 mmol) in DMF (3.0 mL) was stirred

at room temperature for 1 h. Then a solution of 6a (30 mg, 0.043

mmol) and DIPEA (10 µL, 0.043 mmol) in DMF (2.0 mL) was

added and stirring was continued at room temperature overnight.

At the end of the reaction (HPLC control), the solvent was distilled

off and the residue was dissolved in TFA/CH₂Cl₂(1:1; 3.0 mL).

At the end of the reaction (HPLC control), the solvents were

distilled off and the crude product was purified by preparative HPLC

1

51 (24 mg, 61%) as the trifluoroacetate salt. H NMR (400 MHz,

DMSO-d₆): δ 8.30 (1H, s), 8.07 (1H, d), 8.02-7.94 (1H, m), 7.91-

7.76 (4H, m), 5.74 (2H, s), 3.67-3.50 (8H, m), 3.34-3.26 (2H,

m), 3.07 (9H, s), 2.92 (3H, s), 2.68 (3H, s), 2.91 (3H, s), 2.43-

2.36 (2H, m), 2.12-1.33 (14H, m). MS m/z calcd for C₃₇H₅₀-

Cl₂N₅O₅S, 746.29; found, 746.2 [M]⁺. HPLC purity: System B,

99%, t_R) 9.99 min.

1

to obtain 55 (28 mg, 61%) as the trifluoroacetate salt. H NMR

(400 MHz, DMSO-d₆): δ 8.62 (1H, s), 8.04 (1H, d), 7.90-7.32

(12H, m), 5.59 (2H, s), 3.58-3.41 (8H, m), 2.86-2.56 (9H, m),

2.03-1.21 (12H, m). MS m/z calcd for C₃₄H₄₄Cl₂N₆O₅S, 718.25.72;

found, 719.4 [M + H]⁺. HPLC purity: System B, 99%, t_R) 8.79

min.

[(S)-1-(4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxym-

ethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-piperazine-

1-carbonyl)-5-guanidino-pentyl]-trimethyl-ammonium Triflu-

oroacetate Salt (52). A solution of 54 (6.0 mg, 0.005 mmol) in

CH₂Cl₂(2.0 mL) was treated with Goodman’s reagent (8.0 mg,

0.02 mmol) and DIPEA (0.04 mmol) and stirred at room temper-

ature for 1 h. At the end of the reaction (HPLC control), TFA was

added (0.5 mL) and stirring was continued until deprotection of

Boc groups was complete (HPLC control). The solvents were

distilled off, and the residue was purified by preparative HPLC to

[(S)-4-Amino-6-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-

piperazin-1-yl)-6-oxo-hexyl]-trimethyl-ammonium Trifluoroac-

etate Salt (56). A solution of acid 4a (65 mg, 0.124 mmol), HOAt

(22 mg, 0.16 mmol), and EDAC (32 mg, 0.17 mmol) in DMF (2

mL) was stirred in an ice bath for 30 min, then a solution of 21

(58 mg, 0.12 mmol) in DMF (2 mL) was added, followed by

dropwise addition of DIPEA (42 µL, 0.24 mmol). Stirring was

continued at 0 °C for 30 min, then at room temperature overnight.

At the end of the reaction (HPLC control) the solvents were

removed in vacuo. The residue was dissolved in CH₃CN/water

(1:1) and purified by preparative HPLC to obtain the Boc-protected

intermediate as trifluoroacetate salt. This compound was dissolved

in TFA/CH₂Cl₂(1:1; 4 mL) and stirred at room temperature

overnight. The solvents were distilled off in vacuo, and the resulting

product was purified by preparative HPLC to afford 56 (44 mg,

56%) as the trifluoroacetate salt. ¹H NMR (400 MHz, DMSO-d₆):

δ 8.27 (1H, s), 8.08-7.99 (1H, m), 7.95-7.72 (4H, m), 7.59-

7.51 (1H, m), 7.47-7.37 (2H, m), 5.68 (2H, m), 3.76-3.48 (8H,

m), 3.37-3.24 (2H, m), 3.13-3.06 (9H, m), 2.92-2.79 (1H, m),

2.76-2.63 (7H, m), 2.13-1.99 (2H, m), 1.93-1.74 (3H, m), 1.73-

1.60 (2H, m), 1.56-1.42 (4H, m). MS m/z calcd for C₃₇H₅₁-

Cl₂N₆O₅S, 761.30; found, 761.2 [M]⁺. HPLC purity: System B,

98.6%, t_R) 10.26 min.

{(S)-4-Amino-1-[2-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quino-

lin-8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbo-

nyl}-piperazin-1-yl)-2-oxo-ethyl]-butyl}-trimethyl-ammonium

Trifluoroacetate Salt (57). A solution of acid 4a (58 mg, 0.11

mmol), HOAt (19 mg, 0.14 mmol), and EDAC (32 mg, 0.17 mmol)

in DMF (2.0 mL) was stirred in an ice bath for 30 min. Then a

solution of amine 26 (39 mg, 0.073 mmol) in DMF (2.0 mL) was

added while stirring, followed by the dropwise addition of DIPEA

(13 µL, 0.075 mmol). Stirring was continued at 0 °C for an

additional 30 min, then at room temperature overnight. At the end

of the reaction (HPLC control), the solvents were removed in vacuo,

and the residue was dissolved in CH₃CN/water + 0.1% TFA (1:1;

6.0 mL) and purified by preparative HPLC to obtain the Cbz-

protected derivative (36.9 mg, 45%). A portion of this product (14.9

mg, 0.0133 mmol) was dissolved in TFA (1.5 mL), and triflic acid

1

obtain 52 (5.0 mg, 87%) as trifluoroacetate salt. H NMR (400

MHz, DMSO-d₆): δ 8.39 (1H, s), 8.02 (1H, d), 7.78 (1H, d), 7.76

(1H, d), 7.58 (1H, m), 7.35-7.30 (3H, m), 6.95 (4H, br s), 5.65

(2H, s), 4.61 (1H, d), 3.85-3.62 (10H, m), 3.05 (9H, s), 2.72 (3H,

s), 2.68 (3H, s), 1.80-1.69 (3H, m), 1.60-1.53 (7H, m), 1.48-

1.40 (4H, m). MS m/z calcd for C₃₈H₅₃Cl₂N₈O₅S, 803.32; found,

803.3 [M + H]⁺. HPLC purity: System B, 97%, t_R) 7.84 min.

N-[1-[4-(2-(S)-trimethylammonium-6-trimethylammonium-

hexanoyl)-piperazin-1-yl]-cyclopentyl]-2,4-dichloro-3-(2,4-di-

methyl-quinolin-8-yloxymethyl)-benzenesulfonamide Trifluo-

roacetate Salt (53). A solution of acid 36 (10 mg, 0.04 mmol),

EDAC (5.0 mg, 0.04 mmol), and HOAt (8.0 mg, 0.04 mmol) in

DMF (5.0 mL) was cooled in an ice bath. After 1 h, amine 6a (30

mg, 0.046 mmol) and DIPEA (7 µL, 0.046 mmol) were added and

stirring was continued overnight at room temperature. At the end

of the reaction (HPLC control), the solvents were distilled off and

the crude residue was purified by preparative HPLC to obtain 53

1

(20.5 mg, 39%) as the trifluoroacetate salt. H NMR (400 MHz,

DMSO-d₆): δ 8.32 (1H, s), 8.02 (1H, d), 7.78 (1H, d), 7.73 (1H,

d), 7.59-7.31 (3H, m), 5.68 (2H, s), 4.68-4,60 (1H, m), 4.01-

3.56 (8H, m), 3.36-3.28 (2H, m), 3.22 (9H, s), 3.08 (9H, s), 2.68

(3H, s), 2.63 (3H, s), 2.13-1.43 (14H, m). MS m/z calcd for C₄₀H₅₈-

Cl₂N₆O₅S, 804.3; found, 803.3 [M - H]⁺. HPLC purity: System

B, 99.6%, t_R) 8.80 min.

[(S)-5-Amino-1-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-

piperazine-1-carbonyl)-pentyl]-trimethyl-ammonium Trifluo-

roacetateSalt(54).Asolutionofcrude(S)-(5-tert-butoxycarbonylamino-

1-carboxy-pentyl)-trimethyl-ammonium (50 mg, 0.17 mmol), EDAC

(32 mg, 0.17 mmol), and HOAt (23 mg, 0.17 mmol) in DMF (5

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 563

(15 µL, 0.17 mmol) was added. At the end of the reaction (HPLC

control), water (4.0 mL) was added, the resulting mixture was

filtered, and the filtrate was purified by preparative HPLC to obtain

57 (15 mg, quantitative) as the trifluoroacetate salt. ¹H NMR (400

MHz, DMSO-d₆): δ 8.25 (1H, s), 8.02 (1H, d), 7.79-7.69 (5H,

m), 7.56-7.47 (1H, m), 7.44-7.32 (2H, m), 5.67 (2H, s), 3.74-

3.54 (8H, m), 3.23-3.16 (1H, m), 3.09 (9H, s), 2.93-2.71 (4H,

m), 2.68 (3H, s), 2.64 (3H, s), 2.14-2.00 (3H, m), 1.83-1.73 (2H,

m), 1.72-1.55 (3H, m), 1.53-1.41 (4H, m). MS m/z calcd for

C₃₇H₅₁Cl₂N₆O₅S, 761.30; found, 761.3 [M]⁺. HPLC purity: System

B, 97.9%, t_R) 10.02 min.

mmol) and DIPEA (11 µL, 0.04 mmol) were added and stirring

was continued overnight at room temperature. At the end of the

reaction (HPLC control), the solvents were distilled off in vacuo

and the residue was dissolved in CH₂Cl₂(2.0 mL) and TFA (0.5

mL). After complete removal of the Boc group, the solvents were

distilled off and the crude product was purified by preparative HPLC

to obtain 61 (5 mg, 12%) as the trifluoroacetate salt. ¹H NMR (400

MHz, DMSO-d₆): δ 8.61 (1H, s), 8.32 (3H, br s), 8.02 (1H, d),

7.78 (1H, d), 7.81-7.35 (3H, m), 5.62 (2H, s), 4.58 (1H, br s),

3.85 (8H, m), 3.15 (9H, s), 2.72 (3H, s), 2.68 (3H, s), 2.22 (2H,

m), 1.80 (2H, m), 1.60 (2H, m). MS m/z calcd for C₃₅H₄₇Cl₂N₆O₅S,

733.27; found, 733.2 [M]⁺. HPLC purity: System B, 97%, t_R

)

N-{1-[4-((S)-3-Amino-6-guanidino-hexanoyl)-piperazine-1-

carbonyl]-cyclopentyl}-2,4-dichloro-3-(2,4-dimethyl-quinolin-8-

yloxymethyl)-benzenesulfonamide Trifluoroacetate Salt (58). A

solution of acid 17 (18 mg, 0.036 mmol) in DMF (2.0 mL) was

cooled in an ice bath. EDAC (8.0 mg, 0.036 mmol) and HOAt

(5.0 mg, 0.036 mmol) were added, and stirring was continued for

1 h. Amine 6a (20 mg, 0.024 mmol) and DIPEA (9.0 µL, 0.048

mmol) were added, and the resulting mixture was stirred at room

temperature overnight. At the end of the reaction (HPLC control),

the solvents were distilled off in vacuo, and the residue was

dissolved in CH₂Cl₂(2.0 mL) and TFA (0.5 mL). After 12 h, the

solvents were distilled off and the residue was purified by

preparative HPLC to obtain 58 (6.0 mg, 23%) as the trifluoroacetate

8.58 min.

[5-(4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl}-piperazin-1-yl)-

5-oxo-pentyl]-trimethyl-ammonium Trifluoroacetate Salt (62).

A solution of (4-carboxybutyl)-trimethyl ammonium (7.0 mg, 0.04

mmol), EDAC (5.0 mg, 0.04 mmol), and HOAt (8.0 mg, 0.04

mmol) in DMF (5.0 mL) was stirred in an ice bath. After 1 h, amine

6a (20 mg, 0.03 mmol) and DIPEA (8 µL, 0.03 mmol) were added

and stirring was continued overnight at room temperature. At the

end of the reaction (HPLC control), the solvents were distilled off

in vacuo and the crude product was purified by preparative HPLC

1

to obtain 62 (20 mg, 69%) as the trifluoroacetate salt. H NMR

1

salt. H NMR (400 MHz, DMSO-d₆): δ 8.44 (1H, s), 8.02 (1H,

(400 MHz, DMSO-d₆): δ 8.39 (1H, s), 8.03 (1H, d), 7.78 (1H, d),

7.78-7.76 (1H, m), 7.60 (1H, t), 7.54-7.44 (2H, m), 5.65 (2H, s),

3.62 (4H, br s), 3.54 (4H, br s), 3.33-3.27 (2H, m), 3.05 (9H, s),

2.72 (3H, s), 2.68 (3H, s), 2.43 (2H, m), 2.06-1.99 (2H, m), 1.80-

1.69 (4H, m), 1.60-1.53 (2H, m), 1.48-1.40 (4H, m). MS m/z

calcd for C₃₆H₄₈Cl₂N₅O₅S, 732.27; found, 732.4 [M]⁺. HPLC

purity: System B, 97.2%, t_R) 9.27 min.

d), 7.79 (1H, d), 7.77-7.67 (3H, m), 7.50 (1H, t), 7.43 (1H, t),

7.37 (1H, d), 7.32 (1H, br s), 7.10-6.90 (4H, br s), 5.61 (2H, s),

3.77-3.41 (9H, m), 3.02 (2H, m), 2.79-2.68 (2H, m), 2.66 (3H,

s), 2.59 (3H, s), 2.06-1.37 (12H, m). MS m/z calcd for C₃₅H₄₆-

Cl₂N₈O₅S, 760.27; found, 761.3 [M + H]⁺. HPLC purity: System

B, 91%, t_R) 9.02 min.

[2-(4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl}-piperazin-1-yl)-

2-oxo-ethyl]-trimethyl-ammonium Trifluoroacetate Salt (59). A

solution of carboxymethyl-trimethyl-ammonium chloride (6.24 mg,

0.06 mmol), EDAC (15.2 mg, 0.17 mmol), and HOAt (23 mg, 0.17

mmol) in DMF (5.0 mL) was stirred in an ice bath. After 1 h, amine

6a (30 mg, 0.04 mmol) and DIPEA (11 µL, 0.04 mmol) were added

and stirring was continued overnight at room temperature. At the

end of the reaction (HPLC control), the solvent was distilled off in

vacuo and the crude product was purified by preparative HPLC to

obtain 59 (27 mg, 73%) as the trifluoroacetate salt. ¹H NMR (400

MHz, DMSO-d₆): δ 8.27 (1H, s), 8.03 (1H, d), 7.77 (1H, d), 7.74-

7.37 (4H, m), 5.69 (2H, s), 4.51 (2H, s), 3.75-3.47 (8H, m), 3.29

(9H, s), 2.70 (3H, s), 2.66 (3H, s), 2.11-2.01 (2H, m), 1.84-1.73

(2H, m), 1.53-1.43 (4H, m). MS m/z calcd for C₃₂H₄₂Cl₂N₅O₅S,

690.23; found, 690.1 [M]⁺. HPLC purity: System B, 98.1%, t_R)

9.64 min.

[(S)-4-Amino-5-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-

piperazin-1-yl)-5-oxo-pentyl]-trimethyl-ammonium Trifluoro-

acetate Salt (63). A solution of 10b (19 mg, 0.07 mmol), EDAC

(13 mg, 0.07 mmol), and HOAt (9.5 mg, 0.07 mmol) in DMF (5.0

mL) was stirred in an ice bath. After 1 h, amine 6a (34 mg, 0.05

mmol) and DIPEA (24 µL, 0.09 mmol) were added and stirring

was continued overnight at room temperature. At the end of the

reaction (HPLC control), the solvents were distilled off in vacuo

and the residue was dissolved in CH₂Cl₂(2.0 mL) and TFA (0.5

mL). When removal of the Boc group was complete, the solvents

were distilled off and the crude product was purified by preparative

1

HPLC to obtain 63 (15 mg, 27%) as the trifluoroacetate salt. H

NMR (400 MHz, DMSO-d₆): δ 8.33 (1H, s), 8.21 (3H, br s), 8.15

(1H, d), 7.78 (1H, d), 7.75 (1H, d), 7.58-7.50 (1H, t), 7.47-7.35

(2H, m), 5.64 (2H, s), 4.45 (1H, br s), 3.75 (8H, br s), 3.15 (9H,

s), 2.72 (3H, s), 2.68 (2H, s), 1.77 (4H, m), 1.45 (4H, m). MS m/z

calcd for C₃₆H₄₉Cl₂N₆O₅S, 747.8; found, 747.3 [M]⁺. HPLC

purity: System B, 97%, t_R) 8.37 min.

[4-(4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl}-piperazin-1-yl)-

4-oxo-butyl]-trimethyl-ammonium Trifluoroacetate Salt (60). A

solution of (3-carboxypropyl)trimethylammonium chloride (10 mg,

0.068 mmol), EDAC (16 mg, 0.085 mmol), and HOAt (10 mg,

0.073 mmol) in DMF (5.0 mL) was stirred in an ice bath. After 1

h, amine 6a (30 mg, 0.04 mmol) and DIPEA (14 µL, 0.05 mmol)

were added and stirring was continued overnight at room temper-

ature. At the end of the reaction (HPLC control), the solvents were

distilled off in vacuo and the crude product was purified by

preparative HPLC to obtain 60 (21 mg, 55%) as the trifluoroacetate

[5-(4-(1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl]-piperazin-1-yl)-

4(S)-trimethylammonio-5-oxo-pentyl]trimethyl-ammonium Tri-

fluoroacetate Salt (64). A solution of 14 (9.0 mg, 0.04 mmol),

EDAC (5.0 mg, 0.04 mmol), and HOAt (8.0 mg, 0.04 mmol) in

DMF (5.0 mL) was stirred in an ice bath. After 1 h, amine 6a (20

mg, 0.03 mmol) and DIPEA (8 µL, 0.03 mmol) were added and

stirring was continued overnight at room temperature. At the end

of the reaction (HPLC control), the solvents were distilled off in

vacuo and the crude product was purified by preparative HPLC to

obtain 64 (9.0 mg, 26%) as the trifluoroacetate salt. ¹H NMR (400

MHz, DMSO-d₆): δ 8.43 (1H, s), 8.03-8.00 (1H, d), 7.81-7.79

(1H, d), 7.75-7.73 (1H, d), 7.54 (1H, t), 7.44-7.37 (2H, m), 5.64

(2H, s), 4.65 (1H, d), 3.34 (2H, m), 3.20 (6H, s), 3.01 (9H, s), 2.81

(3H, s), 2.73 (3H, s), 2.68 (3H, s), 2.01 (2H, m), 1.77 (2H, m),

1.47 (4H, m). MS m/z calcd for C₃₅H₄₇Cl₂N₆O₅S, 790.34; found,

777.5 [M - CH₃]⁺. HPLC purity: System B, >99%, t_R) 8.55

min.

1

salt. H NMR (400 MHz, DMSO-d₆): δ 8.25 (1H, s), 8.02 (1H,

d), 7.82-7.70 (2H, m), 7.58-7.33 (3H, m), 5.68 (2H, s), 3.64 (4H,

br s), 3.55 (4H, br s), 3.37-3.28 (2H, m), 3.10 (9H, s), 2.69 (3H,

s), 2.65 (3H, s), 2.50-2.44 (2H, m), 2.11-1.73 (6H, m), 1.55-

1.42 (4H, br s). MS m/z calcd for C₃₅H₄₆Cl₂N₅O₅S, 718.24; found,

718.2 [M]⁺. HPLC purity: System B, 97.6%, t_R) 9.71 min.

[(S)-3-Amino-4-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-

piperazin-1-yl)-4-oxo-butyl]-trimethyl-ammonium Trifluoroac-

etate Salt (61). A solution of 10a (13 mg, 0.05 mmol), EDAC

(10 mg, 0.05 mmol), and HOAt (7.0 mg, 0.05 mmol) in DMF (5.0

mL) was stirred in an ice bath. After 1 h, amine 6a (25 mg, 0.04

[(S)-4-Amino-1-(4-{1-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-cyclopentanecarbonyl}-

piperazine-1-carbonyl)-butyl]-trimethyl-ammonium Trifluoro-

564 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3

Fattori et al.

acetate Salt (65). A solution of 12 (11 mg, 0.04 mmol), EDAC

(6.0 mg, 0.04 mmol), and HOAt (8.0 mg, 0.04 mmol) in DMF

(5.0 mL) was stirred in an ice bath. After 1 h, amine 6a (30 mg,

0.04 mmol) and DIPEA (11 µL 0.04 mmol) were added and stirring

was continued overnight at room temperature. At the end of the

reaction (HPLC control), the solvents were distilled off in vacuo

and the residue was dissolved in CH₂Cl₂(2.0 mL) and TFA (0.5

mL). When removal of the Boc group was complete, the solvents

were distilled off and the crude product was purified by preparative

[(S)-4-Amino-5-(4-{4-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-piperidine-4-carbonyl}-

piperazin-1-yl)-5-oxo-pentyl]-trimethyl-ammonium Trifluoro-

acetate Salt (69). A solution of the acid 10b (10 mg, 0.036 mmol),

EDAC (26 mg, 0.036 mmol), and HOAt (6.0 mg, 0.044 mmol) in

DMF (3.0 mL) was stirred at room temperature for 1 h. Amine 6e

(26 mg, 0.036 mmol) and DIPEA (15 µL, 0.088 mmol) in DMF

(1.0 mL) were added and stirring was continued for 1.5 h. At the

end of the reaction (HPLC control), the solvents were distilled off

in vacuo and the residue was dissolved in TFA/CH₂Cl₂(1:1; 4.0

mL) and stirred at room temperature for 40 min. The solvents were

distilled off in vacuo, and the residue was purified by preparative

HPLC to afford 69 (14.9 mg) as the trifluoroacetate salt. ¹H NMR

(400 MHz, DMSO-d₆): δ 9.20 (1H, s), 8.50 (2H, s), 8.25 (3H, s),

8.10-7.30 (6H, m), 5.60 (1H, s), 4.55 (1H, s), 4.00-3.30 (8H,

m), 3.30-2.90 (13H, m), 2.80-2.55 (9H, m), 2.25-1.70 (8H, m).

MS m/z calcd for C₃₆H₅₀Cl₂N₇O₅S, 762.29; found, 762.3 [M]⁺.

HPLC purity: System A, >99%, t_R) 5.17 min.

1

HPLC to obtain 65 (18 mg, 41%) as the trifluoroacetate salt. H

NMR (400 MHz, DMSO-d₆): δ 8.43 (1H, s), 8.03 (1H, d), 7.80-

7.78 (2H, d), 7.74-7.72 (2H, d), 7.54 (1H, t), 7.43-7.38 (2H, m),

5.64 (2H, s), 4.69 (2H, d), 3.20 (9H, s), 2.85 (2H, s), 2.68 (3H, s),

2.63 (3H, s), 2.00 (2H, m), 1.76 (2H, m), 1.46 (4H, m). MS m/z

calcd for C₃₆H₄₉Cl₂N₆O₅S, 747.28; found, 747.3 [M]⁺. HPLC

purity: System B, 98.7%, t_R) 8.64 min.

4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl}-N-(3-dimethyl-

amino-propyl)-piperazine-1-carboxamidine Trifluoroacetate Salt

(66). A solution of acid 4a (78 mg, 0.15 mmol), HOAt (20 mg,

0.15 mmol), and EDAC (28.5 mg, 0.15 mmol) in DMF (3.0 mL)

was stirred at room temperature for 25 min. Then a solution of

amine 38 (40 mg, 0.124 mmol) and DIPEA (65 µL, 0.37 mmol) in

DMF (2.0 mL) was added and stirring was continued at room

temperature overnight. At the end of the reaction (HPLC control),

the solution was concentrated to 2.5 mL and purified by HPLC to

[(S)-4-Amino-5-(4-{4-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-1-methyl-piperidine-4-

carbonyl}-piperazin-1-yl)-5-oxo-pentyl]-trimethyl-ammonium Tri-

fluoroacetate Salt (70). Crude 6e (114 mg, 0.072 mmol) was

dissolved in dry DMF (1 mL), and DIPEA was added until the pH

was about 7. Acid 10 (22 mg, 0.079 mmol), HOAt (10.8 mg, 0.079

mmol), EDAC (15.2 mg, 0.079 mmol), and DIPEA (13.5 µL, 0.079

mmol) were added, and the mixture was stirred at room temperature

overnight. The solvents were distilled off under reduced pressure,

the residue was dissolved in dry DCM (1.5 mL), and TFA (1.5

mL) was added dropwise while stirring, After 30 min, the solvents

were distilled off and the residue was purified by preparative HPLC

1

give 66 (80 mg, 50%) as the trifluoroacetate salt. H NMR (400

MHz, DMSO-d₆): δ 9.78-9.40 (1H, br s), 8.35-8.22 (1H, m),

8.06-7.94 (1H, m), 7.82-7.57 (5H, m), 7.57-7.46 (1H, m), 7.46-

7.32 (2H, m), 5.71-5.63 (2H, m), 3.80-3.66 (4H, m), 3.59-3.48

(4H, m), 3.36-3.26 (2H, m), 3.15-3.06 (2H, m), 2.86-2.78 (6H,

m), 2.73-2.58 (6H, m), 2.12-1.98 (2H, m), 1.98-1.87 (2H, m),

1.83-1.72 (2H, m), 1.54-1.41 (4H, m). MS m/z calcd for C₃₄H₄₅-

Cl₂N₇O₄S, 717.26; found, 718.2 [M + H]⁺. HPLC purity: System

B, 93.4%, t_R) 10.14 min.

{3-[(4-{1-[2,4-Dichloro-3-(2,4-dimethyl-quinolin-8-yloxymethyl)-

benzenesulfonylamino]-cyclopentanecarbonyl}-piperazine-1-

carboximidoyl)-amino]-propyl}-trimethyl-ammonium Trifluo-

roacetate Salt (67). An ice-cold solution of 4a (121 mg, 0.23

mmol), HOAt (31 mg, 0.23 mmol), and EDAC (44 mg, 0.23 mmol)

in DMF (5.0 mL) was stirred for 20 min. Then a solution of the

amine 40 (65 mg, 0.19 mmol) and DIPEA (19 µL, 0.57 mmol) in

DMF (2.0 mL) was added. At the end of the reaction, the solvents

were distilled off in vacuo and the crude product was purified by

preparative HPLC to afford 67 (7.5 mg, 4%) as the trifluoroacetate

salt. ¹H NMR (400 MHz, DMSO-d₆): δ 8.37-8.26 (1H, m), 8.07-

7.98 (1H, m), 7.83-7.66 (4H, m), 7.56-7.46 (1H, m), 7.44-7.31

(2H, m), 5.73-5.64 (2H, m), 3.82-3.71 (4H, m), 3.62-3.52 (5H,

m), 3.41-3.26 (4H, m), 3.18-3.06 (9H, m), 2.74-2.60 (6H, m),

2.14-1.96 (5H, m), 1.85-1.73 (2H, m), 1.56-1.43 (4H, m). MS

m/z calcd for C₃₅H₄₈Cl₂N₇O₄S, 732.29; found, 732.1 [M]⁺. HPLC

purity: System B, 98.5%, t_R) 9.90 min.

1

to obtain 70 (15 mg, 21%) as the trifluoroacetate salt. H NMR

(400 MHz, DMSO-d₆, 353 °K): δ 8.90 (1H, s), 8.19 (2H, br s),

8.05 (1H, d, J ) 8.7 Hz), 7.80 (1H, d, J ) 8.7 Hz), 7.74 (1H, d,

J ) 8.3 Hz), 7.52 (1H, t, J ) 8.1 Hz), 7.41 (1H, d, J ) 7.7 Hz),

7.33 (1H, s), 5.66 (2H, s), 4.47 (1H, s), 4.13-3.38 (12H, covered

by water), 3.38-3.27 (2H, m), 3.07 (9H, s), 2.69-2.64 (6H, m),

2.61 (3H, s), 2.38-1.95 (4H, m), 1.89-1.70 (4H, m). MS m/z calcd

for C₃₇H₅₂Cl₂N₇O₅S, 776.31; found, 776.3 [M]⁺. HPLC purity:

System A, >99%, t_R) 8.20 min.

[(S)-5-(4-{1-Acetyl-4-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-piperidine-4-carbonyl}-

piperazin-1-yl)-4-amino-5-oxo-pentyl]-trimethyl-ammonium Tri-

fluoroacetate Salt (71). A solution of acid 10 (8.0 mg, 0.026

mmol), HOAt (4.0 mg, 0.026 mmol), EDAC (5.6 mg, 0.026 mmol),

and DIPEA (4.5 µL, 0.026 mmol) in DMF (2 mL) was stirred at

room temperature for 1 h and then added to crude 6f (18 mg, 0.025

mmol) dissolved in dry DMF (1 mL). At the end of the reaction

(HPLC control), the solvents were distilled off under reduced

pressure and the residue was dissolved in DCM/TFA (1:1; 2 mL)

and stirred for 2 h. The solvents were distilled off, and the crude

product was purified by preparative HPLC to obtain 71 (10 mg,

1

38%) as the trifluoroacetate salt. H NMR (400 MHz, DMSO-d₆,

353 °K): δ 8.55 (1H, s), 8.15 (2H, br s), 8.02 (1H, d, J ) 8.7 Hz),

7.77 (1H, d, J ) 8.7 Hz), 7.70 (1H, d, J ) 8.2 Hz), 7.47 (1H, t,

J ) 8.2 Hz), 7.36-7.29 (2H, m), 5.67 (2H, s), 4.46 (1H, s), 3.93-

3.56 (8H, m), 3.55-3.06 (6H, m, covered by water), 3.07 (9H, s),

2.65 (3H, s), 2.61 (3H, s), 1.89 (3H, s), 1.85-1.71 (8H, m). MS

m/z calcd for C₃₈H₅₂Cl₂N₇O₆S, 804.3; found, 804.2 [M]⁺. HPLC

purity: System B, 97%, t_R) 14.23 min.

[(S)-5-Amino-6-(4-{4-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-tetrahydro-pyran-4-car-

bonyl}-piperazin-1-yl)-6-oxo-pentyl]-trimethyl-ammonium Hy-

drochloride Salt (72). A solution of the acid 10 (75 mg, 0.27

mmol), EDAC (62 mg, 0.32 mmol), and HOAt (44 mg, 0.32 mmol)

in DMF (3.0 mL) was stirred at room temperature for 1 h. Amine

6b (150 mg, 0.18 mmol) and DIPEA (110 µL, 0.6 mmol) in DMF

(1.0 mL) were added, and stirring was continued for 20 h. At the

end of the reaction (HPLC control), the solvents were distilled off

in vacuo and the residue was dissolved in CH₂Cl₂, washed with

5% NaHCO₃and satd NaCl, dried over Na₂SO₄, and concentrated

under reduced pressure. The residual oil was dissolved in a small

[(S)-4-Amino-5-(4-{4-[2,4-dichloro-3-(2,4-dimethyl-quinolin-

8-yloxymethyl)-benzenesulfonylamino]-tetrahydro-thiopyran-4-

carbonyl}-piperazin-1-yl)-5-oxo-pentyl]-trimethyl-ammonium Tri-

fluoroacetate Salt (68). A solution of the acid 10b (36 mg, 0.13

mmol), EDAC (75 mg, 0.39 mmol), and HOAt (18 mg, 0.13 mmol)

in DMF (3.0 mL) was stirred at room temperature for 1 h. Then a

solution of amine 6d (86 mg, 0.13 mmol) and DIPEA (44 µL, 0.26

mmol) in DMF (1.0 mL) was added and stirring was continued for

1 h. At the end of the reaction (HPLC control), the solvents were

distilled off in vacuo and the residue was dissolved in TFA (2.0

mL) and stirred at room temperature for 40 min. Et₂O was added

with stirring and a product was precipitated that was filtered off.

Purification by preparative HPLC afforded 68 (14 mg, 11%) as

1

the trifluoroacetate salt. H NMR (400 MHz, DMSO-d₆): δ 8.80

(1H, s), 8.0-7.40 (6H, m), 5.60 (1H, s), 4.55 (1H, s), 3.80-3.30

(8H, m), 3.0 (9H, s), 2.70 6H, s), 2.40-2.00 (8H, m), 1.75 (4H,

d). MS m/z calcd for C₃₆H₄₉Cl₂N₆O₅S₂, 779.25; found, 779.2 [M]⁺.

HPLC purity: System A, >99%, t_R) 7.52 min.

Bradykinin hB₂Receptor Antagonists

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 3 565

(6) Svensson, C.; Andresson, M.; Persson, C. G.; Venge, P.; Alkner,

U.; Pipkorn, U. Albumin, bradykinins, and eosinophil cationic protein

on the nasal mucosal surface in patients with hay fever during natural

allergen exposure. J. Allergy Clin. Immunol. 1990, 85, 828-833.

(7) Akbary, A. M.; Wirth, K. J.; Scholkens, B. A. Efficacy and tolerability

of Icatibant (Hoe 140) in patients with moderately severe chronic

bronchial asthma. Immunopharmacology 1996, 33, 238-242.

(8) Austin, C. E.; Foreman, J. C.; Scadding, G. K. Reduction by Hoe

140, the B₂kinin receptor antagonist, of antigen-induced nasal

blockage. Br. J. Pharmacol. 1994, 111, 969-971.

(9) Kuoppala, A.; Lindsted, K. A.; Saarinen, J.; Kovanen, P. T.;

Kokkonen, J. O. Inactivation of bradykinin by angiotensin-converting

enzyme and by carboxypeptidase N in human plasma. Am. J. Physiol.

2000, 278 (4), H1069-1074.

(10) Campbell, D. J.; Krum, H.; Esler, M. D. Losartan increases bradykinin

levels in hypertensive humans. Circulation 2005, 111, 315-320.

(11) Fattori, D.; Rossi, C.; Fincham, C. I.; Berettoni, M.; Calvani, F.;

Catrambone, F.; Felicetti, P.; Gensini, M.; Terracciano, R.; Altamura,

M.; Bressan, A.; Giuliani, S.; Maggi, C. A.; Meini, S.; Valenti, C.;

Quartara, L. Design and synthesis of novel sulfonamide-containing

bradykinin hB₂receptor antagonists. 1. Synthesis and SAR of R,R-

dimethylglycine sulfonamides. J. Med. Chem. 2006, 49, 3602-3613.

(12) Sum of residual BK-induced response calculated in percentage with

respect to the basal response and normalized in comparison to the

control registered at the various times of observation (5-30-60-

90-120-180-210 min), further calculated in percentages with

respect to the theoretical maximum. As a consequence, the maximal

inhibition that may be obtained corresponds to Σ% ) 0, while the

absence of inhibition corresponds to Σ% ) 100.

(13) Cucchi, P.; Meini, S.; Bressan, A.; Catalani, C.; Bellucci, F.;

Santicioli, P.; Lecci, A.; Faiella, A.; Rotondaro, L.; Giuliani, S.;

Giolitti, A.; Quartara, L.; Maggi, C. A. MEN16132, a novel potent

and selective nonpeptide antagonist for the human bradykinin B2

receptor. In vitro pharmacology and molecular characterization. Eur.

J. Pharmacol. 2005, 528, 7-16.

volume of CH₂Cl₂(1.0 mL) and precipitated with Et₂O. Filtration

gave the crude Boc-protected derivative (145 mg, 0.17 mmol, 94%).

This was dissolved in CH₂Cl₂(3.0 mL), and 4 N HCl in dioxane

(3.0 mL) was added. After stirring at room temperature for 30 min,

the solvents were removed in vacuo. Trituration of the residue with

Et₂O afforded 72 (MEN 16132; 110 mg, 73%) as the hydrochloride

salt. ¹H NMR (400 MHz, DMSO-d₆): δ 8.85 (1H, s), 8.50 (3H, s),

8.02 (1H, d), 7.90-7.60 (4H, m), 5.59 (2H, s), 4.57-4.45 (1H,

m), 3.70-3.18 (12H, m), 3.08 (9H, s), 2.81 (6H, m), 1.95-1.60

(8H, m). MS m/z calcd for C₃₆H₄₉Cl₂N₆O₆S, 763.2; found, 763.1

[M]⁺. HPLC purity: System B, 97%, t_R) 7.40 min.

(B) Biology. The receptor binding assays and the measurement

of inositol monophosphate accumulation were carried out as

described in ref 11. The measurement of the antagonist potency

(pA₂) in the BK-induced contraction of the GPI was performed as

described in ref 13.

In vivo experiments were performed in male Dunkin Hartley

guinea pigs weighing 350-400 g (Charles River, Italy) in ac-

cordance with the European Union and the local ethical commitee

regulations. Evaluation of bronchoconstriction and hypotension

induced by iv bradykinin and the antagonist effects of the

compounds were performed according to the methods reported in

ref 16.

Acknowledgment. We are grateful to Mr. Ricci and Mr.

Fabbri for technical assistance, to Dr. A. Cartoni, Dr. A. Triolo,

and Dr. S. Mauro for analytical assistance, and to Dr. A. Ettorre

and Dr. G. Balacco for the NMR spectra. This work was

supported by grants from the Italian Ministry of University and

Research (RIF.506/DSPAR 98).

References

(1) Mak, J. C. W.; Barnes, P. J. Autoradiographic visualization of

bradykinin receptors in human and guinea-pig lung. Eur. J. Phar-

macol. 1991, 194, 37-43.

(14) (a) McAllistar, S. M.; Alpar, H. O.; Teitelbaum, Z.; Bennet, D. B.

Do interactions with phospholipids contribute to the prolonged

retention of polypeptides within lungs? AdV. Drug DeliVery ReV.

1996, 19, 89-110. (b) Upton, R. N.; Doolette, D. J. Kinetic aspects

of drug disposition in the lungs. Clin. Exp. Pharmacol. Physiol. 1999,

26, 381-391.

(2) Dear, J. W.; Wirth, K.; Scadding, G. K.; Foreman, J. C. Characteriza-

tion of the bradykinin receptor in the human nasal airway using the

binding of [¹²⁵I]-Hoe 140. Br. J. Pharmacol. 1996, 119, 1054-1062.

(3) Polosa, R.; Holgate, S. T. Comparative airway response to inhaled

bradykinin, kallidin, and [des-Arg⁹]bradykinin in normal and asth-

matic subjects. Eur. J. Pharmacol. 1989, 163, 195-197.

(4) Churchill, L.; Pongracic, J. A.; Reynolds, C. J.; Naclerio, R. M.;

Proud, D. Pharmacology of nasal provocation with bradykinin:

Studies of tachyphylaxis, cyclooxygenase inhibition, R-adrenergic

stimulation and receptor subtype. Int. Arch. Allergy Appl. Immunol.

1991, 95, 322-331.

(15) Barnes, P. G. Tiotropium Bromide. Exp. Opin. InV. Drugs 2001, 10,

733-740.

(16) Valenti, C.; Cialdai, C.; Giuliani, S.; Lecci, A.; Tramontana, M.;

Meini, S.; Quartara, L.; Maggi, C. A. MEN 16132, a novel, potent

and selective non-peptide kinin B₂receptor antagonist. In vivo activity

on bradykinin-induced bronchoconstriction and nasal mucosa mi-

crovascular leakage in anesthetized guinea pigs. J. Pharmacol. Exp.

Ther. 2005, 616-623.

(5) Shibayama, Y; Skoner, D.; Suehiro, S.; Konishi, J. E.; Fireman, P.;

Kaplan, A. P. Bradykinin levels during experimental nasal infection

with rhinovirus and attenuated influenza virus. Immunopharmacology

1996, 33, 311-313.

JM061143K

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Design and synthesis of novel sulfonamide-containing bradykinin hB 2 receptor antagonists. 2. Synthesis and structure-activity relationships of α,α-cycloalkylglycine sulfonamides

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