Specific Chymase Inhibitor Research Articles

Attenuation of adhesion formation after cardiac surgery with a chymase inhibitor in a hamster model

Objective Chymase is one of the inflammatory mediators and is released from mast cells, which are closely associated with adhesion formation. Chymase also activates transforming growth factor β1, which promotes tissue fibrosis. However, the role of chymase in cardiac adhesion formation has not yet been elucidated. We have assessed whether a specific chymase inhibitor, Suc-Val-Pro-Phe p (OPh) 2, prevents postoperative cardiac adhesions in hamsters. Methods In 66 hamsters the epicardium was abraded, and then either chymase inhibitor or placebo was injected into the left thoracic cavity, leaving the pericardium open. Cardiac chymase activity, the level of transforming growth factor β1 in the pleural fluid, and the density of epicardial mast cells were measured 3 days postoperatively. The degree of adhesion formation was evaluated macroscopically and histologically 2 weeks postoperatively by using a grading score ranging from 0 (no adhesions) to 4 (severe adhesions). Results The cardiac chymase activity and level of transforming growth factor β1 were lower in the chymase inhibitor–treated group compared with in the placebo-treated group (45.8 ± 18.7 vs 79.7 ± 13.7 μU/mg protein [ P < .025] and 15.6 ± 6.5 vs 33.2 ± 9.8 μg/mL [ P < .01], respectively). The density of mast cells was higher in the placebo-treated group, and there was suppression to 60% of this value in the chymase inhibitor–treated group. The adhesion scores were lower in the chymase inhibitor–treated group compared with in the placebo-treated group (1.3 ± 1.3 vs 3.0 ± 1.1, P < .01). Conclusion Use of a chymase inhibitor suppresses not only cardiac chymase activity but also the level of transforming growth factor β1, and this results in a reduction in postoperative cardiac adhesion.

Chymase-derived angiotensin II and arrhythmias after myocardial infarction.

Mast cell-derived chymase seems to be important in the regulation of local angiotensin (A) II formation in cardiovascular tissues. In human heart, chymase accounts for 80% of A II formation. Therefore, the chymase-dependent A II pathway may play an important role in the pathogenesis of A II-related cardiovascular diseases. For example, cardiac chymase was activated earlier than ACE and this activation lasted longer than that of ACE after myocardial infarction (MI) in hamsters. Treatment with a specific chymase inhibitor treatment, but not an ACE inhibitor, improved post-MI survival as well as cardiac function and the extent of the beneficial effects was similar to that observed for an AT1-receptor antagonist treatment in this model. The survival benefit after MI seems to be related to an antiarrhythmic effect of the chymase inhibitor because chymase inhibition reduces the incidence of ventricular arrhythmias during periods of heart ischemia in a dog MI model. Thus, an antiarrhythmic effect of the chymase inhibitor may contribute to a reduction in mortality rate during the acute phase after MI.

Significance of chymase inhibition for prevention of adhesion formation

To clarify the role of chymase in adhesion formation, we investigated whether a chymase inhibitor could prevent adhesion formation after surgery in hamsters. Hamsters received a lesion produced by uterus scraping. A specific chymase inhibitor, 2-[4-(5-fluoro-3-methylbenzo[ b]thiophen-2-yl)sulfonamido-3-methanesulfonylphenyl]oxazole-4-carboxylicacid (TY-51184), or placebo was injected into the abdomen before closing and scores for adhesion formation were assessed at 1, 4, and 12 weeks. A single peritoneal administration of TY-51184 significantly decreased the adhesion scores even at 12 weeks (placebo, 2.80±0.20; chymase inhibitor, 1.60±0.31). Thus, chymase inhibitors may be a novel strategy to prevent adhesion formation.

Impact of chymase inhibitor on cardiac function and survival after myocardial infarction.

Recent studies have demonstrated that hamsters, like humans, possess both angiotensin converting enzyme (ACE)- and chymase-dependent angiotensin (Ang) II-forming pathways in cardiovascular tissues. We recently found that, after myocardial infarction (MI) in hamsters, cardiac chymase was significantly activated. In order to determine whether suppression of cardiac chymase activity could provide prognostic benefit after MI, we examined the effects of NK3201, a novel, orally active and specific chymase inhibitor, on cardiac function and survival during the acute phase of MI in hamsters. Two hundred and ten male Syrian hamsters were used in the present study. The left coronary artery of each hamster was ligated to induce MI. NK3201 at a dose of 30 mg/kg per day was administered orally by gastric gavage, starting either 3 days before or 1 day after MI. ACE and chymase activities were significantly increased in the infarcted left ventricle 3 days after MI. NK3201 treatment starting 3 days before MI significantly inhibited the increase in cardiac chymase activity, while it did not affect ACE activity either in plasma or in heart 3 days after MI. A significant improvement in cardiac function was observed 3 and 14 days after MI in the group receiving NK3201. Compared with vehicle treatment, NK3201 treatment initiated either 3 days before or 1 day after MI significantly reduced the mortality rate during the 14 days of observation following MI. These findings indicate that cardiac chymase plays an important role after MI and this finding may provide a novel therapeutic target in post-MI treatment.

Cardiac mast cells in the transition to heart failure

Cardiac mast cells in the transition to heart failure

Chymase inhibition prevents cardiac fibrosis and improves diastolic dysfunction in the progression of heart failure.

Angiotensin (Ang) II, which plays a crucial role in the cardiac remodeling process, is generated via angiotensin-converting enzyme (ACE); however, an alternative generation pathway, chymase, which is stored in the mast cells, also exists in the heart. Cardiac chymase is insensitive to ACE inhibitors (ACEIs), and heart chymase promotes interstitial fibrosis by affecting collagen metabolism via transforming growth factor-beta in vitro. Therefore, selective chymase blockade seems to be an important strategy in the prevention of cardiac remodeling We evaluated the effects of a specific chymase inhibitor, SUNC8257 (Chy I; 10 mg/kg twice a day; n=7), on changes in cardiac structures, Ang II levels, and gene expressions, which are characterized as molecular markers for fibrosis, in dogs with tachycardia induced heart failure (HF). In HF, the number of chymase enzyme-positive mast cells increased in the left ventricle (LV) compared with the normal group; however, Chy I significantly decreased the mast cell density and cardiac Ang II levels. Despite no significant differences in LV systolic function compared with the vehicle group, Chy I decreased LV end-diastolic pressure and shortened the prolongation of tau. Chy I suppressed collagen-type I and III and transforming growth factor-beta mRNA levels and decreased fibrosis in the LV compared with the vehicle. The chymase pathway may be critical for cardiac diastolic dysfunction accompanied with fibrosis. Chronic chymase inhibition may therefore become an important strategy in the prevention of cardiac remodeling in HF.

Suppression of basic fibroblast growth factor-induced angiogenesis by a specific chymase inhibitor, BCEAB, through the chymase-angiotensin-dependent pathway in hamster sponge granulomas.

1. We investigated the profound involvement of mast cell chymase, an alternative angiotensin II-generating enzyme, in angiogenesis using a specific chymase inhibitor. We also studied the functional profiles of this novel inhibitor in basic fibroblast growth factor (bFGF)-induced angiogenesis. 2. In this study, angiogenesis was induced by daily injections of bFGF (0.3 micro g site(-1) day(-1)), angiotensin I (2 nmol site(-1) day(-1)) or angiotensin II (2 nmol site(-1) day(-1)) into sponges implanted to male hamsters subcutaneously for 7 days. Angiogenesis in the granulation tissue surrounding sponges was evaluated by measuring the haemoglobin (Hb) content and local blood flow as the parameters for angiogenesis. 3. A chymase inhibitor, BCEAB (4-[1-[[bis-(4-methyl-phenyl)-methyl]-carbamoyl]-3-(2-ethoxy-benzyl)-4-oxo-azetidine-2-yloxy]-benzoic acid), was simultaneously administered into the implanted sponges (2 or 5 nmol site(-1) day(-1), for 7 days) treated with bFGF and strongly suppressed the haemoglobin contents in sponge granulomas. In the studies using a laser doppler perfusion imager, BCEAB (5 nmol site(-1) day(-1)) also attenuated the bFGF-induced increase of local blood flow around the implanted sponge granuloma. 4. In bFGF-induced angiogenesis, chymase activity in sponge granulomas was substantially increased. It was also confirmed that the chymase activity increased by bFGF was significantly and dose-dependently inhibited by BCEAB (2, 5 nmol site(-1) day(-1)). 5. BCEAB inhibited the Hb contents and the expression of vascular endothelial growth factor (VEGF) mRNA induced by angiotensin I but not by angiotensin II. 6. These results suggest that the significance of chymase in bFGF-induced angiogenesis was confirmed, and a novel inhibitor, BCEAB, strongly suppresses the bFGF-induced angiogenesis through the chymase-angiotensin II-VEGF dependent pathway.

Chymase Inhibitor, BCEAB, Suppressed Peritoneal Adhesion Formation in Hamster

Background. Mast cells are closely related to adhesion formation, while it has been unclear which factor in mast cells plays an important role in the development of adhesion formation. To clarify the role of chymase produced from mast cells in adhesion formation, we investigated the preventive effect of a specific chymase inhibitor, BCEAB, on adhesion formation in a hamster experimental model. Materials and methods. Hamsters were administered orally once daily with 100 mg/kg of BCEAB or placebo from the operated day to 1 week after the operation. The uterus was grasped and denuded by a swab. Results. One week after the operation, the scores for adhesion formation in the chymase inhibitor-treated group were significantly decreased in comparison with those in the placebo-treated group (placebo-treated group, 2.80 ± 0.20; chymase inhibitor-treated group 1.60 ± 0.31: P < 0.01). The chymase activity in the injured uterus was also significantly suppressed in the chymase inhibitor-treated group (placebo-treated group, 17.3 ± 2.69 mU/mg protein; chymase inhibitor-treated group 9.60 ± 0.89: P < 0.05). After scraping the utelus, the level of transforming growth factor-β in the peritoneal fluid was significantly increased in the placebo-treated group, while it was suppressed to 70% by the treatment with BCEAB. Conclusions. The specific chymase inhibitor BCEAB may be a useful drug for prevention of adhesion formation.

Beneficial effects of cardiac chymase inhibition during the acute phase of myocardial infarction

Recently, the presence of the chymase-dependent angiotensin (Ang) II-generating system in hamsters, dogs, monkeys, as well as human cardiovascular tissues has been identified. We have reported that the activation of cardiac chymase was more prominent than that of angiotensin converting enzyme (ACE) and that AT1 receptor antagonist treatment rather than ACE inhibitor treatment alone provided significant beneficial effects on cardiac function and survival after MI in hamsters. The aim of the present study was to determine whether this different effects between AT1 receptor antagonist and ACE inhibitor were due to the activation of cardiac chymase after MI in hamsters by using 4-[1-{[bis-(4-methyl-pheny)-methyl]-carbamoyl}-3-(2-ethoxy-benzyl)-4-oxo-azetidine-2-yloxy]-benzoic acid (BCEAB), a novel, orally active and specific chymase inhibitor. The ACE and chymase activities in the infarcted left ventricle were significantly increased 3 days after MI. BCEAB (100 mg/kg/day, p.o.) treatment starting 3 days before MI significantly suppressed the cardiac chymase activity, while it did not affect the plasma and cardiac ACE activities 3 days after MI. A significant improvement in hemodynamics (maximal negative and positive rates of pressure development; left ventricular systolic pressure) was observed for the treatment with BCEAB 3 days after MI. BCEAB (100 mg/kg/day, p.o.) treatment starting 3 days before MI significantly reduced the mortality rate during 14 days of observation following MI (vehicle, 61.1%, n = 18; BCEAB, 27.8%, n = 18; P < 0.05). These findings demonstrated for the first time that cardiac chymase participates directly in the pathophysiologic state after MI in hamsters.

Development and Application of Chymase Inhibitors: Therapeutic Potential of a Specific Chymase Inhibitor in Atopic Dermatitis

A novel therapeutic mechanism may be the key to improving the chief symptoms and signs of atopic dermatitis (AD), which are persistent pruritus and high serum IgE. We demonstrate here that mast cell chymase may be a possible initiating factor and that the orally active specific inhibitor Y-40613 may have a therapeutic potential in the treatment of AD. We found that Y-40613 (2-[5-amino-2-(4-fluorophenyl)-1,6-dihydro-6-oxo-1-pyrimidinyl]-N-[1-[(5-methoxycarbonyl-2-benzoxazolyl)carbonyl]-2-phenylethyl]acetamide) dose-dependently suppressed the scratching response in a mouse pruritus model, with inhibitory efficacy enhanced by combination with conventional drugs, suggesting that chymase contributes to the development of pruritus by a unique mechanism or mechanisms. In fact, chymase injected in the model induced the scratching response. In vitro IgE production from mouse B cells was increased by purified rat chymase and suppressed by Y-40613. Increased serum IgE observed in Brown Norway rats injected with mercury chloride was suppressed by Y-40613. Furthermore, Y-40613 lowered ear thickness as well as serum IgE level in a mouse contact dermatitis model. Taken together, these findings suggest that the specific chymase inhibitor Y-40613 may ameliorate symptoms of AD through the dual inhibition of the chymase-dependent IgE production pathway and itching sensation.

Chymase inhibitor suppresses adhesion formation in a hamster experimental model

To clarify the role of chymase produced by mast cells in adhesion formation, we investigated the preventive effect of a specific chymase inhibitor, Suc-Val-Pro-Phe p (OPh) 2, on adhesion formation in a hamster experimental model. Hamsters underwent resection of the right uterine body and then 10 μM Suc-Val-Pro-Phe p (OPh) 2 or placebo was injected into the abdomen. Two weeks after the operation, the scores for adhesion formation in the chymase inhibitor-treated group were significantly lower than that in the placebo-treated group (placebo-treated group, 3.60±0.22; chymase inhibitor-treated group, 2.10±0.22; P<0.01). This specific chymase inhibitor, Suc-Val-Pro-Phe p (OPh) 2, significantly suppressed the scores for adhesion formation in a hamster experimental model. Thus, chymase may play an important role in the adhesion formation.

Oral administration of a specific chymase inhibitor, NK3201, inhibits vascular proliferation in grafted vein

Chymase may play an important role in vascular proliferation, as shown by in-vitro experiments, but the role of chymase in vivo has been unclear. In this study, we investigated the effect of a novel chymase inhibitor, NK3201, on this proliferation in dog grafted veins. NK3201 inhibited human and dog chymases, but not rabbit ACE. NK3201 suppressed the Ang I-induced vascular contraction in isolated dog arteries in the presence of an ACE inhibitor, and the IC 50 value of chymostatin and NK3201 in dog artery was 320 nM. In dog, the concentration of NK3201 in blood was about 10 μM at 24 h after oral administration of the drug (5 mg/kg). In the group treated with NK3201, each dog was administered orally 5 mg/kg per day from 5 days before to the day before the removal of the grafted veins. Each dog underwent right common carotid artery bypass grafting with the ipsilaterial external jugular vein. By 28 days after grafting, a significant vascular proliferation was observed in the grafted veins and the chymase activity was also increased significantly. Treatment with chymase inhibitor significantly suppressed the proliferation of the grafted veins and the increased chymase activity. In this study, we demonstrate for the first time that oral administration of a specific chymase inhibitor, NK3201, appears useful for preventing vascular proliferation.