BK-induced Bronchoconstriction Research Articles

Design and Synthesis of Novel Sulfonamide-Containing Bradykinin hB2 Receptor Antagonists. 2. Synthesis and Structure−Activity Relationships of α,α-Cycloalkylglycine Sulfonamides

Recently we reported on the design and synthesis of a novel class of selective nonpeptide bradykinin (BK) B2 receptor 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.

Design and Synthesis of Novel Sulfonamide-Containing Bradykinin hB2 Receptor Antagonists. 1. Synthesis and SAR of α,α-Dimethylglycine Sulfonamides

We recently published the extensive in vivo pharmacological characterization of MEN 16132 (J. Pharmacol. Exp. Ther. 2005, 616-623; Eur. J. Pharmacol. 2005, 528, 7), a member of the sulfonamide-containing human B(2) receptor (hB(2)R) antagonists. Here we report, in detail, how this family of compounds was designed, synthesized, and optimized to provide a group of products with subnanomolar affinity for the hB(2)R and high in vivo potency after topical administration to the respiratory tract. The series was designed on the basis of indications from the X-ray structures of the key structural motifs A and B present in known antagonists and is characterized by the presence of an alpha,alpha-dialkyl amino acid. The first lead (17) of the series was submitted to extensive chemical work to elucidate the structural requirements to increase hB(2) receptor affinity and antagonist potency in bioassays expressing the human B(2) receptor (hB(2)R). The following structural features were selected: a 2,4-dimethylquinoline moiety and a piperazine linker acylated with a basic amino acid. The representative lead compound 68 inhibited the specific binding of [(3)H]BK to hB(2)R with a pKi of 9.4 and antagonized the BK-induced inositolphosphate (IP) accumulation in recombinant cell systems expressing the hB(2)R with a pA(2) of 9.1. Moreover, compound 68 when administered (300 nmol/kg) intratracheally in the anesthetized guinea pig, was able to significantly inhibit BK-induced bronchoconstriction for up to 120 min after its administration, while having a lower and shorter lasting effect on hypotension.

MEN16132, a Novel Potent and Selective Nonpeptide Kinin B2Receptor Antagonist: In Vivo Activity on Bradykinin-Induced Bronchoconstriction and Nasal Mucosa Microvascular Leakage in Anesthetized Guinea Pigs

We have tested the activity of 4-(S)-amino-5-(4-[4-[2,4-dichloro-3-(2,4-dimethyl-8-quinolyloxymethyl)phenylsulfonamido]-tetrahydro-2H-4-pyranylcarbonyl] piperazino)-5-oxopentyl](trimethyl)ammonium chloride hydrochloride (MEN16132), a novel nonpeptide kinin B(2) receptor antagonist, on bradykinin (BK)-induced inflammatory responses, bronchoconstriction, and hypotension in guinea pigs. After i.v. (1-10 nmol/kg i.v.), intratracheal (i.t.) (10-100 nmol/kg i.t.), or aerosol (0.01-0.1 mM/5 min) administration, MEN16132 inhibited in a dose-dependent manner the bronchoconstriction induced by BK (10 nmol/kg i.v.). MEN16132 was more potent and possessed a longer duration of action as compared with the peptide B(2) receptor antagonist icatibant (HOE140; H-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-Tic-Oic-Arg-OH trifluoroacetate). After i.v. administration, its inhibitory effect on bronchoconstriction lasted more than 8 h at 30 nmol/kg. When administered by i.v. or i.t. routes, the dose completely inhibiting bronchoconstriction also partially reduced the hypotensive response to BK, whereas after aerosol administration, the inhibitory effect was limited to respiratory level. Intranasal (i.n.) administration of MEN16132 (0.01-0.3 nmol/nostril) reduced, in a dose-dependent and long-lasting manner, the nasal mucosa plasma protein extravasation induced by BK (100 nmol/nostril), and it exerted a complete inhibition at about 30-fold lower dose than icatibant. At 1 nmol/nostril, MEN16132 activity was significant for at least 6 h with no systemic effect measured as inhibition of BK-induced hypotension, and at 10 nmol/nostril, the inhibitory effect lasted for more than 15 h with only a weak effect on hypotension. These findings indicate that in vivo MEN16132 is a potent kinin B(2) receptor antagonist with long duration of action, both after i.v. and local administration. A complete and prolonged inhibition of BK-induced bronchoconstriction or nasal inflammation can be achieved with MEN16132 topical administration (aerosol or i.n.) at doses devoid of systemic effects.

A new class of nonpeptide bradykinin B(2) receptor ligand, incorporating a 4-aminoquinoline framework. Identification of a key pharmacophore to determine species difference and agonist/antagonist profile.

Introduction of various aliphatic amino groups at the 4-position of the quinoline moiety of our nonpeptide bradykinin (BK) B(2) receptor antagonists afforded highly potent ligands for human B(2) receptor with various affinities for guinea pig B(2) receptor, indicating remarkable species difference. A representative 4-dimethyamino derivative 40a exhibited subnanomolar and nanomolar binding affinities for human and guinea pig B(2) receptors, respectively, and significantly inhibited BK-induced bronchoconstriction in guinea pigs at 10 microg/kg by intravenous administration. Further chemical modification led us to discover unique partial agonists for the human B(2) receptor that increase inositol phosphates (IPs) production by themselves in Chinese hamster ovary (CHO) cells expressing the cloned human B(2) receptor. Although their potency and efficacy were much lower than those of BK, we identified them as screening leads for nonpeptide B(2) agonists. In these studies it was revealed the 4-substituent of the quinoline moiety is the key pharmacophore to determine species difference and agonist/antagonist profiles.

A New Series of Highly Potent Non-Peptide Bradykinin B2 Receptor Antagonists Incorporating the 4-Heteroarylquinoline Framework. Improvement of Aqueous Solubility and New Insights into Species Difference

Introduction of nitrogen-containing heteroaromatic groups at the 4-position of the quinoline moiety of our non-peptide B(2) receptor antagonists resulted in enhancing binding affinities for the human B(2) receptor and reducing binding affinities for the guinea pig one, providing new structural insights into species difference. A CoMFA study focused on the diversity of the quinoline moiety afforded correlative and predictive QSAR models of binding for the human B(2) receptor but not for the guinea pig one. A series of 4-(1-imidazolyl)quinoline derivatives could be dissolved in a 5% aqueous solution of citric acid up to a concentration of 10 mg/mL. A representative compound 48a inhibited the specific binding of [(3)H]BK to the cloned human B(2) receptor expressed in Chinese hamster ovary cells with an IC(50) value of 0.26 nM and significantly inhibited BK-induced bronchoconstriction in guinea pigs even at 1 microg/kg by intravenous administration.

In vitro and in vivo activity of analogues of the kinin B2 receptor antagonist MEN1 1270.

In this study, we describe the in vitro and in vivo activities of a series of cyclic peptide analogues of the selective kinin B2 receptor antagonist MEN11270 on Chinese hamster ovary cells expressing the human B2 receptor (hB2R), the human isolated umbilical vein (hUV), the isolated guinea pig ileum (gpI), and bradykinin (BK) induced bronchoconstriction (BC) and hypotension in anaesthetized guinea pigs. Substitutions in the backbone of MEN1 1270 (H-DArg-Arg-Pro-Hyp-Gly-Thi-c(Dab-DTic-Oic-Arg)c(7gamma-10alpha)) aimed to increase the potency in inhibiting bronchospasm versus hypotension following the topical (intratracheal (i.t.)) or systemic (intravenous (i.v.)) application of these antagonists. A series of analogues were left unprotected from N-terminal cleavage by aminopeptidases (MEN12739, MEN13052, MEN13346, and MEN13371): these compounds maintained sizeable affinities for the hB2R (pKi = 9.4, 9.6, 9.7, and 8.6, respectively) and antagonist activities toward BK in the hUV (pA2 = 7.9, 8.3, 8.2, and 7.5) and gpI assays (pK(B) = 7.4, 7.8, 7.9, and 7.9), but the inhibition of BK-induced BC and hypotension in vivo was negligible following either i.v. or i.t. administration. Two analogues (MEN12388 and MEN13405) could be potential substrates of angiotensin-converting enzyme: these have good activity in the hB2R (pKi = 9.5 and 8.9, respectively), hUV (pA2 = 8.2 for MEN12388), and gpI assays (pK(B) = 8.4 and 8.0) but an in vivo activity 10- to 30-fold lower than the parent compound MEN1 1270 (pKi = 9.4, pA2 = 8.1, pKB = 8.3) when given by either the i.v. or the i.t. route. Other analogues were functionalized with a quaternary ammonium Lys derivative (MEN13031, MEN12374, and the previously mentioned MEN13052) or with an ethyl group on Arg (MEN13655 and the previously mentioned MEN13346 and MEN13405) in order to hinder or facilitate local absorption. MEN13346 and MEN13031 (pKi = 9.7and 9.5, pA2 = 8.2 and 7.9, pKB = 7.9 and 8.5, respectively) were 10- to 30-fold less active in vivo than MEN1 1270, without improving the discrimination between BK-induced BC and hypotension after either systemic or topical administration. It is concluded that the decreased in vivo activities of cyclic analogues of MEN11270 on BK-induced BC and hypotension following either their intratracheal or their intravenous routes of administration might be due in large part to metabolic degradation.

Effect of neutral endopeptidase inhibitor on bradykinin-induced bronchoconstriction

To evaluate whether neutral endopeptidase (NEP) inhibitors have adverse respiratory effects, the influence of a NEP inhibitor on bradykinin (BK)-induced bronchoconstriction was investigated. In anesthetized and artificially ventilated guinea pigs, changes in airway opening pressure (Pao) were measured as an index of bronchoconstriction. An infusion of phosphoramidon (3 mg kg −1 h −1), a NEP inhibitor, significantly enhanced the bronchoconstriction induced by high-dose BK (30 nmol kg −1, i.v.). Capsaicin (0.1 mg kg −1, i.v.) and SR48968 (0.3 mg kg −1, i.v.), an NK2 receptor antagonist, significantly inhibited the phosphoramidon-induced enhancement of BK-induced bronchoconstriction, although FK888 (3 mg kg −1, i.v.), an NK1 receptor antagonist, did not. Both neurokinin A (NKA) (0.1–3 nmol kg −1, i.v.) and substance P (SP) (0.1–3 nmol kg −1, i.v.) induced dose-dependent bronchoconstriction which was enhanced by phosphoramidon infusion, although these enhancements were more prominent in the NKA series. Phosphoramidon partially inhibited BK degradation in lung homogenate, and both NKA and SP degradation in the lung homogenate were significantly suppressed by phosphoramidon. In bronchoalveolar lavage fluid (BALF), levels of NKA and SP were significantly elevated after a bolus of BK with a phosphoramidon infusion. These results suggest that NEP inhibitors may have adverse respiratory effects resulting from inhibition of the degradation of neurokinins, but mainly of NKA, when a large amount of BK is generated.

Bradykinin-induced Airway Constriction in Guinea-pigs: Role of Leukotriene D4

Tachykinins (TK) have been implicated in both bradykinin-(BK) and hyperpnea-induced broncho-constriction (HIB) in the guinea-pig. However, TKs appear to have an indirect effect in HIB by releasing leukotriene (LT)D4. We postulated that BK may cause bronchoconstriction through a cascade involving TK and LTD4. We examined the role of TK and LTD4in BK-induced bronchoconstriction in ventilated Hartley guinea-pigs. Respiratory resistance (Rrs) was monitored for 2 h following insufflation of BK (150 nM). Animals were pretreated with propranolol, then with either neurokinin (NK)1 (CP-99,994)+NK2 (SR-48,968) receptor antagonists or pranlukast (90 μ g or 900 μ g), an LTD4antagonist. Control animals received no pretreatment. BK-induced bronchoconstriction was significantly lower in NK1/NK2 (128%±6% baseline RrsSEM) and pranlukast (90 μ g; 205±22, 900μ g; 169±20) animals compared to controls (284±22), P<0.0001 ANOVA. Bile from control and saline challenged animals was analysed for LTD4by HPLC and radio-immunoassay. However, LTD4excretion rate showed no significant difference over a 2-h collection period following insufflation of either BK or saline, respectively; baseline =2.5 pmol/h±0.6 SEM vs. 2.3±0.2, 0–1 h=2.8±0.7 vs. 2.0±0.6, 1–2 h=2.3±0.6 vs. 1.7±0.7. We conclude that BK-induced bronchoconstriction is mediated in part through the release of both TK and LTD4, but the latter is released in insufficient quantities to be detectable by biliary analysis.

Contribution of bradykinin B(1) and B(2) receptors in allergen-induced bronchial hyperresponsiveness.

Bradykinin (BK) is a peptide mediator generated at sites of inflammation and its effects are mediated through constitutively expressed B(2) receptor or through induction of B(1) receptors. We examined the role of these receptors in bronchial hyperresponsiveness (BHR). Brown-Norway rats sensitized with ovalbumin (OA) and Al(OH)(3) intraperitoneally, were exposed 3 wk later to either saline or OA aerosol. B(1) receptor antagonist desArg(10)[Hoe140] (200 nmol/kg or 1 micromol/kg, intraperitoneally) or B(2) receptor antagonist Hoe140 (200 nmol/kg, intraperitoneally) was administered 30 min before allergen exposure. Hoe140 had no effect on OA-induced BHR to acetylcholine (ACh) and bronchoalveolar lavage fluid (BALF) cellular profiles, but inhibited bronchoconstriction to BK (p < 0.04). At both doses, desArg(10)[Hoe140] dose-dependently inhibited allergen-induced BHR to ACh (p < 0.01), but had no effect on bronchoconstriction to BK or baseline ACh responsiveness. The inflammatory cells in BALF were not affected apart from reduced lymphocyte numbers at the highest dose. B(1) receptor mRNA expression measured by Northern analysis was increased after allergen exposure in sensitized lungs, with a peak at 2 to 6 h after exposure, whereas B(2) receptor mRNA expression remained unchanged. Newly induced BK B(1) receptors may be involved in allergen-induced BHR to ACh, whereas constitutive B(2) receptors mediate BK-induced bronchoconstriction.

Discovery of orally active nonpeptide bradykinin B2 receptor antagonists

Orally active nonpeptide bradykinin (BK) B2 receptor antagonists have been discovered by using directed random screening and chemical modification. These compounds displaced [3H]BK binding to B2 receptors in guinea-pig ileum membranes, rat uterus membranes and human lung fibroblasts with nanomolar IC50s. They did not inhibit different specific radio-ligand bindings to other receptor sites including B1 receptors. In isolated guinea-pig ileum preparations, these compounds had no agonistic effect on smooth muscle contraction at 10−6 M, and caused parallel rightward shifts of the concentration–response curves to BK on contraction with higher pA2 values. They also blocked human B2 receptor-mediated phosphatidylinositol hydrolysis without agonistic effect. In vivo, the oral administrations of these antagonists potently inhibited BK-induced bronchoconstriction in guinea-pigs. They also reduced carrageenin-induced paw edema and caerulein-induced pancreatitis in rats. Moreover, these compounds alleviated kaolin-induced pain in mice by oral administration. These results show that our compounds are potent, selective, and orally active BK B2 receptor antagonists and that they may have therapeutic potential against inflammatory diseases and pain.

A novel class of orally active non-peptide bradykinin B2 receptor antagonists. 4. Discovery of novel frameworks mimicking the active conformation.

In recent articles we reported the identification of a series of 8-[[2, 6-dichloro-3-[N-methyl-N-[(E)-(substituted)acryloylglycyl]amino]++ +benzy l]oxy]-2-methylimidazo[1,2-a]pyridines as the first orally active non-peptide bradykinin (BK) B2 receptor antagonists. Optimization of the terminal glycine part and the imidazo[1,2-a]pyridine moiety led to the discovery of a clinical candidate (5, FR173657). With the aim of completion of the structure-activity relationship (SAR) study, we next investigated the roles of the substituents on the central phenyl ring. The results suggested that the 2,6-dichloro or 2, 6-dimethyl groups may play important roles in regulating the conformations of the 1- and 3-substituents and also may interact with hydrophobic pockets of the B2 receptors. Furthermore, according to the results of a molecular modeling study reported in part 1 of this series, we designed and synthesized a series of sterically constrained analogues by replacing the N-methylamide group with cis-amide-like rigid moieties. We discovered several bioisosteres and chemically proved that the N-methylamide moiety adopts the cis-amide form in the active conformation. Extensive chemical modification led to the identification of a novel class of highly potent and orally active non-peptide B2 antagonists represented by a pyrrole derivative (52a, FR193517). Compound 52a inhibited the specific binding of [3H]BK to recombinant human B2 receptors expressed in Chinese hamster ovary (CHO) cells and guinea pig ileum membrane preparations expressing B2 receptors with IC50s of 0.37 and 0.56 nM, respectively. This compound also displayed excellent in vivo functional antagonistic activity against BK-induced bronchoconstriction in guinea pigs at 1 mg/kg by oral administration.

A novel class of orally active non-peptide bradykinin B2 receptor antagonists. 2. Overcoming the species difference between guinea pig and man.

Recently we reported the identification of a series of 8-[[3-(N-acylglycyl-N-methylamino)-2, 6-dichlorobenzyl]oxy]-3-halo-2-methylimidazo[1,2-a]pyridines as the first orally active non-peptide bradykinin (BK) B2 receptor antagonists (1-3). These compounds inhibited the specific binding of [3H]BK to guinea pig ileum membrane preparations expressing B2 receptors with nanomolar IC50's and also displayed in vivo functional antagonistic activities against BK-induced bronchoconstriction in guinea pigs at 1 mg/kg by oral administration. However, it was found that their affinities for the B2 receptors in human A-431 cells (human epidermoid carcinoma) were much lower. Intensive modifications of the terminal substituents at the glycine moiety elucidated the structure-activity relationships (SAR) for human B2 receptors, leading to an extended basic framework which incorporated a novel key pharmacophore. Thus, we overcame the species difference and identified the first clinical candidate 18c (FR167344) with IC50's of 0.66 and 1.4 nM for guinea pig ileum and human A-431 cells, respectively. This compound displayed in vivo functional antagonistic activity against BK-induced bronchoconstriction in guinea pigs with an ED50 value of 0.17 mg/kg by oral administration. This novel non-peptide B2 antagonist is extremely potent both in vitro and in vivo by oral administration and is expected to be the first member of a new class of drug for the treatment of various inflammatory diseases.

A novel class of orally active non-peptide bradykinin B2 receptor antagonists. 3. Discovering bioisosteres of the imidazo[1,2-a] pyridine moiety.

Recently we reported on overcoming the species difference of our first orally active non-peptide bradykinin (BK) B2 receptor antagonists, incorporating an 8-[[3-(N-acylglycyl-N-methylamino)-2, 6-dichlorobenzyl]oxy]-3-halo-2-methylimidazo[1,2-a]pyridine skeleton, leading to identification of the first clinical candidate 4a (FR167344). With this potent new lead compound in hand, we then investigated further refinement of the basic framework by replacement of the imidazo[1,2-a]pyridine moiety and discovered several bioisosteric heterocycles. Extensive optimization of these new heteroaromatic derivatives revealed the detailed structure-activity relationships (SAR) around the imidazo[1, 2-a]pyridine ring and the 2,6-dichlorobenzyl moiety, leading to the discovery of our second clinical candidate 87b (FR173657) which inhibited the specific binding of [3H]BK to recombinant human B2 receptors expressed in Chinese hamster ovary (CHO) cells and guinea pig ileum membrane preparations expressing B2 receptors with IC50's of 1.4 and 0.46 nM, respectively. This compound also displayed excellent in vivo functional antagonistic activity against BK-induced bronchoconstriction in guinea pigs with an ED50 value of 0.075 mg/kg by oral administration. Further modifications of the terminal substituents on the pyridine moiety led to a novel pharmacophore and resulted in the identification of 99 (FR184280), whose IC50 value for human B2 receptors (0.51 nM) was comparable to that of the second-generation peptide B2 antagonist Icatibant.

Inhibitory effect of NPC-17731 on BK-induced and antigen-induced airway reactions in guinea-pigs.

Bradykinin (BK) has been suggested to act as a mediator in the airways in inflammatory conditions, such as asthma through the activation of B2-receptors. NPC-17731 (D-Arg0[Hyp3, D-HypE(trans-propyl)7, Oic8]BK) has potent antagonistic activity against B2-receptors without agonistic activity. We have evaluated the inhibitory effect of NPC-17731 against BK in guinea-pig airways. In addition, we have investigated the effects of NPC-17731 on antigen-induced airway responses. Bronchoconstriction was assessed as an increase in lung resistance (RL) and a decrease in dynamic compliance (Cdyn). Airway plasma leakage was assessed by extravasation of intravenously injected Evans blue dye. To estimate the effect of drugs on antigen-induced reactions, guinea-pigs were actively sensitized by exposure to aerosol ovalbumin (OA) twice and challenged by OA inhalation. Acute bronchoconstriction was measured for 15 min. Airway vascular leakage was measured at 10 min after the challenge. Assessment of airway hyperresponsiveness against acetylcholine and bronchoalveolar lavage were conducted at 18-24 h after the antigen-challenge. NPC-17731 (0.3-30 microg/kg, i.v.) inhibited intravenously applied BK-induced bronchoconstriction in a dose-dependent manner. The 50% inhibitory doses (ID50) were 1.3 microg/kg for RL and 2.8 microg/kg for Cdyn. NPC-17731 (1-10 microg/kg, i.v.) inhibited BK-induced microvascular leakage in a dose-dependent manner (ID50 = 4.2 microg/kg). In addition, 10 microg/kg of NPC-17731 abolished the inhaled BK-induced bronchoconstriction. In the sensitized animals, 100 microg/kg NPC-17731 significantly reduced the airway microvascular leakage and the decrease in Cdyn induced by ovalbumin exposure (P < 0.05), but did not influence the increase in RL. NPC-17731 (100 microg/kg) inhibited the antigen-induced airway hyperresponsiveness and the increase in eosinophils in BAL fluids. These results indicate that NPC-17731 is a potent BK antagonist in vivo and that BK may partially contribute to the antigen-induced airway responses in guinea-pigs.

A novel class of orally active non-peptide bradykinin B2 receptor antagonists. 1. Construction of the basic framework.

A novel class of potent, selective, and orally active non-peptide bradykinin (BK) B2 receptor antagonists were designed and synthesized starting from 8-benzyloxyimidazo[1,2-a]pyridine derivative 2. The unique screening lead (2) was discovered by a two-step intentional random screening process, involving recognition of the relationship between BK and angiotensin II (Ang II) and the common structural features. Systematic chemical modification of 2 elucidated the structural requirements essential for B2 binding affinity leading to the identification of 8-[[3-(N-acylglycyl-N-methylamino)-2,6-dichlorobenzyl]oxy]-3-halo- 2- methylimidazo[1,2-a]pyridine skeleton as the basic framework of this new series of B2 antagonists. A molecular modeling study suggested the key role of the N-methylanilide moiety at the 3-position of the 2,6-dichlorobenzene ring to allow these compounds to adopt the characteristic active conformation. The representative lead compounds inhibited the specific binding of [3H]BK to guinea pig ileum membrane preparations expressing B2 receptors, with nanomolar IC50S and also displayed in vivo functional antagonistic activities against BK-induced bronchoconstriction in guinea pigs at an oral dose of 1 mg/kg. Pharmacokinetic studies of compounds 47c and 50b in rats highlighted their excellent oral bioavailabilities, indicating that they represent the first orally active non-peptide B2 antagonists reported to date.

Role of tachykinins in enhancement of bradykinin-induced bronchoconstriction by captopril.

In anesthetized, mechanically ventilated guinea pigs, infusion of captopril (1 mg/kg/h), an angiotensin converting enzyme inhibitor, significantly enhanced bronchoconstriction induced by intravenous injection of bradykinin (BK; 0.1-30 nmol/kg). Pretreatment of guinea pigs with capsaicin (100 mu g/kg) slightly suppressed the bronchoconstriction by BK alone and almost all of the enhancement of BK-induced bronchoconstriction by captopril was suppressed. Intravenous injection of substance P (SP; 0.1-100 nmol/kg), neurokinin A (NKA; 0.1-30 nmol/kg) and neurokinin B (NKB; 0.1-30 nmol/kg) also induced dose-dependent bronchoconstriction but captopril treatment enhanced only the bronchoconstriction induced by SP. BK degradation in bronchoalveolar lavage fluid (BALF) in vitro was significantly suppressed by captopril (p < 0.05). Captopril infusion to guinea pigs significantly increased the levels of BK, SP, and NKA in BALF after BK injection (p < 0.05). FK224, an NK1 and NK2 receptor antagonist and SR 48968, an NK2 receptor antagonist, significantly suppressed the bronchoconstriction induced by BK alone (p < 0.01 and p < 0.05, respectively) as well as the enhancement by captopril (p < 0.01). It can be concluded that the enhancement of BK-induced bronchoconstriction by captopril was attributable to inhibition of the degradation of BK itself and thereby enhanced release of NKA and partly of SP from sensory nerves by BK.

Hoe 140 a new potent and long acting bradykinin-antagonist: in vivo studies.

1. The potency, duration of action and tolerability of Hoe 140, a novel and highly potent bradykinin (BK) antagonist in vitro, has been tested in different in vivo models and compared with the well-known BK antagonist D-Arg-[Hyp2, Thi5,8, D-Phe7]BK. 2. Hoe 140 is highly potent and long acting in inhibiting BK-induced hypotensive responses in the rat. Four hours after s.c. administration of 20 nmol kg-1, inhibition still amounted to 60% whereas the effect of 200 nmol kg-1 of D-Arg-[Hyp2, Thi5,8, D-Phe7]BK was not significant. 3. BK-induced bronchoconstriction in guinea-pigs was strongly inhibited by Hoe 140. The magnitude and duration of inhibition confirmed the findings obtained in the blood pressure experiments in the rat. 4. Carrageenin-induced inflammatory oedema of the rat paw was considerably inhibited at i.v. doses between 0.1 and 1 mg kg-1. 5. In conscious dogs, intravenous doses of 0.01 and 0.1 mg kg-1 of Hoe 140 and D-Arg-[Hyp2, Thi5,8, D-Phe7]BK were well tolerated. At doses of 1 mg kg-1 adverse effects occurred that were attributed to the residual BK agonistic activity of both compounds. 6. Hoe 140 has been shown to be a highly potent and long acting BK antagonist in vivo in different animal species and models. This makes it appropriate to investigate further the physiological and pathophysiological role of BK.

Airway effects of inhaled bradykinin, substance P, and neurokinin A in sheep

The effects of inhaled bradykinin (BK), substance P (SP), and neurokinin A (NKA) on pulmonary resistance and airway responsiveness to carbachol were studied in conscious allergic sheep. Inhaled BK (20 breaths, 0.1 to 5.0 mg · ml −1) caused dose-dependent increases in pulmonary resistance. Neither inhaled SP nor NKA (20 breaths, 0.1 to 1.0 mg · ml −1) produced significant bronchoconstriction in allergic sheep. However, the response to SP could be enhanced ( p < 0.05) by pretreatment with the neutral endopeptidase inhibitor, thiorphan (40 breaths, 1 mg · ml −1). Sheep that were allergic to Ascaris suum antigen were 5.9 times ( p < 0.05) more sensitive to the constrictor effects of BK than nonallergic sheep. BK-induced bronchoconstriction was blocked in a dose-dependent fashion by the BK β 2-receptor antagonist, NPC 567 ( d-arginine{hydroxyproline 3, d-phenylalanine 7}BK). Atropine (0.2 mg · kg −1, intravenously) and nedocromil sodium (1 mg · kg −1 in 3 ml of saline, aerosolized) significantly inhibited the BK-induced bronchoconstriction by 97% and 43%, respectively. Chlorpheniramine (2 mg · kg −1, intravenously) had no effect. NKA caused a transient increase in airway responsiveness in allergic sheep, producing a mean 1.9-fold leftward shift in dose-response curves to aerosolized carbachol ( p < 0.05). This hyperresponsiveness was not evident 24 hours after NKA challenge. Neither SP nor BK changed airway responsiveness. Thus, in allergic sheep, inhaled BK caused a more pronounced bronchoconstriction than that observed in nonallergic sheep. The bronchoconstriction was blocked by a BK-receptor antagonist and appeared to be partially mediated via cholinergic reflexes. The sensory neuropeptides, unlike BK, were not potent bronchoconstrictors when they were administered by inhalation, but NKA may play a role in modulating airway responsiveness.

Bradykinin-induced Airway Microvascular Leakage and Bronchoconstriction Are Mediated via a Bradykinin B2Receptor

We have investigated the effects of bradykinin (Bk) antagonists on Bk-induced airway microvascular leakage and bronchoconstriction in mechanically ventilated guinea pigs by simultaneous measuring extravasation of Evans blue dye and airway opening pressure (Pao). Bradykinin (1 microgram/kg intravenously) significantly increased leakage of dye in trachea, main bronchi, and intrapulmonary airways, and increased Pao, indicating airway narrowing. The selective Bk B2 antagonist D-Arg-[Hyp3-Thi5,8-D-Phe7]-Bk (NPC 349; 400 micrograms/kg intravenously) did not alter basal leakage, but when given 2 min before Bk, significantly inhibited the Bk-induced plasma extravasation by 99.8% in the trachea (p less than 0.05), by 75.9% in main bronchi (p less than 0.05), by 83.5% in proximal intrapulmonary airways (p less than 0.05), and by 91.5% in distal intrapulmonary airways (p less than 0.05). NPC 349 also reduced the increase in Pao induced by Bk by 73.5% (p less than 0.05) without affecting the basal Pao. However, NPC 349 had no inhibitory effect on Bk-induced leak or bronchoconstriction when given 30 min before Bk. By contrast, the B1 antagonist des Arg9-Leu8-Bk (500 micrograms/kg intraveneously) had no effect on the Bk-increased plasma extravasation and Pao. We conclude that Bk increases airway plasma leakage and bronchoconstriction via activation of B2 receptors.