Effect of ABT-627 (A-147627), a potent selective ET(A) receptor antagonist, on the cardiopulmonary profile of newborn lambs with surgically-induced diaphragmatic hernia.

1. Postnatal mortality in isolated congenital diaphragmatic hernia (CDH) is mainly related to the associated pulmonary hypertension (PH) and to right-to-left shunting. 2. Endothelins (ETs) are potent vasoconstrictors and pro-mitogenic peptides. Strong evidences support their participation in CDH and in the etiology of PH via the activation of ET(A) receptors (ET(A)-Rs). 3. Evaluation of the effect of ABT-627, a selective non-peptidic ET(A)-R antagonist, given from -15 to 210 min post-delivery (1 mg kg(-1) bolus +0.01 mg kg(-1) h(-1) infusion, i.v.), was conducted in the lamb model of CDH. 4. Severity of CDH was assessed in comparison to untreated controls (n=5). Untreated CDH lambs (n=7) had a higher mean pulmonary arterial pressure (MPAP; P<0.0001), lower mean blood pressure (MBP; P=0.0004), higher MPAP / MBP ratio (P<0.0001), lower arterial pH (P<0.0001), higher paCO(2) (P<0.0001), lower paO(2) (P<0.0001) and lower post-ductal pulsatile SaO(2) (P<0.0001) than untreated controls. 5. Treated controls (n=7) showed a higher MPAP, lower MBP, higher MPAP/MBP ratio, lower arterial pH, higher paCO(2), lower paO(2), lower post-ductal pulsatile SaO(2) and lower plasmatic ir-ET ratios compared to untreated controls (P<0.0001). 6. Treated CDH lambs (n=8) showed a higher MBP (P<0.0001), lower MPAP / MBP ratio (P<0.0001), higher arterial pH (P<0.0001), lower paCO(2) (P<0.0001), higher paO(2) (P=0.0228), higher post-ductal pulsatile SaO(2) (P=0.0016) and lower plasmatic ir-ET ratios (P=0.0247) when compared to untreated CDH lambs. 7. These observations revealed that, although acute perinatal treatment with a selective non-peptidic ET(A)-R antagonist had some adverse effects in controls, it attenuated the progressive cardiopulmonary deterioration that occurred after birth in CDH lambs.

Pyrrolidine-3-carboxylic acids as endothelin antagonists. 5. Highly selective, potent, and orally active ET(A) antagonists.

The synthesis and structure-activity relationships (SAR) of a series of pyrrolidine-3-carboxylic acids as endothelin antagonists are described. The data shows an increase in selectivity when the methoxy of Atrasentan (ABT-627) is replaced with methyl, and the benzodioxole is replaced with dihydrobenzofuran. Adding a fluorine further increases the binding activity and provides a metabolically stable and orally bioavailable ET(A)-selective antagonist.

'Irreversible' endothelin-1 binding does not prohibit ABT-627 from reversing endothelin-1-induced effects.

Endothelin-1 (ET-1) is thought to play a role in a wide range of pathological conditions. One of the distinct characteristics of ET-1 is its long-lasting vasoconstrictor action, which is presumably caused by the irreversible binding of ET-1 to ET receptors and by the functional effects of internalized ET receptors. ABT-627 is a potent endothelin-A (ET(A))-selective antagonist with a Ki value at 0.034 nM for the human ET(A) receptor, and is currently being used in clinical studies for prostate cancer. Unlike ET-1, the binding of 125I-labeled ABT-627 to human ET(A) receptors expressed in Chinese hamster ovary (CHO) cells is reversible, and the dissociation half-life for the ligand/receptor complex is 2 h. Interestingly, the binding of both ET-1 and ABT-627 to the ET(A)-receptor results in partial receptor internalization but only ET-1 is capable of triggering intracellular functional responses. Although ABT-627 binding to membranes is more reversible than ET-1 binding, ABT-627 is able to reverse an ET-1-induced contraction in rat aortic rings in a dose-dependent manner, and at 1 microM produces nearly complete reversal of the constrictor effects of 10 nM ET-1 within 60 min. Similarly, in vivo studies show that ABT-627 (0.01 and 0.1 mg/kg/min i.v.) reverses the ET-1-induced increase in arterial pressure in anesthetized, ganglionic-blocked rats, and after 60 min, ABT-627 essentially normalizes pressure. Our data show that ABT-627 is capable of reversing an established response induced by ET-1 and is useful in reversing pathological conditions involving ET-1.

Increased susceptibility to deoxycorticosterone acetate-salt-induced hypertension in endothelin-B-receptor-deficient rats.

We evaluated the role of endothelin-B- (ET(B)) receptor-mediated action in the development and maintenance of deoxycorticosterone acetate (DOCA)-salt-induced hypertension, cardiovascular hypertrophy and renal damage, using the spotting lethal (sl) rat which carries a naturally occurring deletion in the ET(B)-receptor gene. Homozygous (sl/sl) rats exhibit abnormal development of the neural crest-derived epidermal melanocytes and the enteric nervous system (ENS), and do not live beyond 1 month because of intestinal aganglionosis and resulting intestinal obstruction. Therefore, the dopamine-beta-hydroxylase (D betaH) promoter was used to direct ET(B) transgene expression in sl/sl rats to support normal ENS development. D betaH-ET(B) sl/sl rats live into adulthood and are healthy, expressing ET(B)-receptor in adrenals and other adrenergic neurons. When homozygous (sl/sl) and wild-type (WT) (+/+) rats, all of which were transgenic, were treated with DOCA and salt for 4 weeks, the homozygous rats exhibited significantly earlier and higher increases in systolic blood pressure than WT rats. The daily oral administration of ABT-627, a selective ET(A)-receptor antagonist, almost completely suppressed the DOCA-salt-induced hypertension in both groups. Renal dysfunction and histological damage induced by DOCA-salt treatment were more severe in homozygous than in WT rats. Increased and marked vascular hypertrophy of the aorta was also observed in homozygous rats, compared with WT rats. Renal and vascular injuries induced by DOCA and salt were significantly improved by ABT-627 administration. We propose that ET(B)-receptor-mediated actions are protective factors in the pathogenesis of DOCA-salt-induced hypertension. ET(A)-mediated actions are at least partly responsible for the increased susceptibility to DOCA-salt-induced hypertension and related tissue injuries in ET(B)-receptor-deficient rats.

Effects of endothelin receptor antagonists on the plasma immunoreactive endothelin-1 level.

Endothelin (ET) receptor antagonists may be beneficial for treating several medical conditions. Human trials with various ET receptor antagonists show that these antagonists elevate the plasma immunoreactive endothelin-1 (irET-1) level, and different classes of antagonists seem to affect the plasma ET-1 level differently. In this report, we study effects of ETA-selective, ETB-selective, and nonselective receptor antagonists on the plasma irET-1 level in the rat, and also compare available clinical data. The plasma irET-1 level was increased by five- and ten-fold after rats were treated with A-192621, an ETB-selective antagonist with Ki values for ETA and ETB at 5600 and 8.8 nM, for 3 days at 30 and 100 mg/kg/day via food. The plasma irET-1 level was increased by 1.8 and 2.4-fold when rats were treated with A-216546, an antagonist with Ki values for ETA and ETB at 0.46 and 13 000 nM, at 10 and 50 mg/kg/day via food for 7 days. As a comparison, the plasma irET-1 level was increased by > 24-fold when rats were treated with A-182086, a nonselective antagonist with Ki values for ETA and ETB at 0.2 and 1.2 nM, at 100 mg/kg/day via food for 9 days. In humans, blockade of ETA by ABT-627 did not result in an elevation in irET-1 until after 7 days of treatment. The results are consistent with the hypothesis that the ETB-receptor is the clearance receptor for ET-1. Our data also suggest that the modest effect of ETA antagonists on the plasma irET-1 level is probably a result of the upregulation of the ET-1 gene via a feedback mechanism.

Exaggerated vascular and renal pathology in endothelin-B receptor-deficient rats with deoxycorticosterone acetate-salt hypertension.

BACKGROUND: Endothelin (ET)-1 plays an important role in the pathogenesis of deoxycorticosterone acetate (DOCA)-salt-induced hypertension. We evaluated the pathological role of ET(B) receptors in DOCA-salt-induced hypertension, cardiovascular hypertrophy, and renal damage by using the spotting-lethal (sl) rat, which carries a naturally occurring deletion in the ET(B) receptor gene. METHODS AND RESULTS: Homozygous (sl/sl) rats exhibit abnormal development of neural crest-derived epidermal melanocytes and the enteric nervous system, and they do not live beyond 1 month because of intestinal aganglionosis and intestinal obstruction. The dopamine ss-hydroxylase (DssH) promoter was used to direct ET(B) transgene expression in sl/sl rats to support normal enteric nervous system development. DssH-ET(B) sl/sl rats live into adulthood and are healthy, expressing ET(B) receptors in adrenal glands and other adrenergic neurons. When homozygous (sl/sl) and wild-type (+/+) rats, all of which were transgenic, were treated with DOCA-salt, homozygous rats exhibited earlier and higher increases in systolic blood pressure than did wild-type rats. Chronic treatment with ABT-627, an ET(A) receptor antagonist, completely suppressed DOCA-salt-induced hypertension in both groups. Renal dysfunction and histological damage were more severe in homozygous than in wild-type rats. Marked vascular hypertrophy was observed in homozygous rats than in wild-type rats. Renal and vascular injuries were significantly improved by ABT-627. In DOCA-salt-treated homozygous rats, there were notable increases in renal, urinary, and aortic ET-1, all of which were normalized by ABT-627. CONCLUSIONS: ET(B)-mediated actions are protective in the pathogenesis of DOCA-salt-induced hypertension. Enhanced ET-1 production and ET(A)-mediated actions are responsible for the increased susceptibility to DOCA-salt hypertension and tissue injuries in ET(B) receptor-deficient rats.

Endothelin and ET(A) receptors in long-term arterial pressure homeostasis in conscious nonhuman primates.

This study was designed to quantify the long-term contribution of endogenous endothelin-1 (ET-1) and ET(A) receptors to the regulation of arterial pressure under normal conditions in nonhuman primates. Therefore, mean arterial pressure (MAP) and heart rate were measured 24 h/day with the use of telemetry techniques in conscious cynomolgus monkeys under control conditions, during administration of an ET(A) selective receptor antagonist (ABT-627; 5 mg/kg, 2 times a day by mouth, 4 days), and a 6-day posttreatment period. Systemic ET(A) blockade reduced MAP (24 h) from 89 +/- 3 to 82 +/- 2 and 79 +/- 2 mmHg on days 1 and 4, respectively. Subsequently, MAP remained suppressed for 3 days posttreatment. Heart rate increased from 111 +/- 5 to 122 +/- 4 and 128 +/- 6 beats/min on days 1 and 4 of ABT-627, respectively, and remained above control for 3 days posttreatment. Plasma ET-1 concentration increased from 1.0 +/- 0.3 to 1.9 +/- 0.4 pg/ml in response to ABT-627 (day 4) but decreased to control values 4 days posttreatment. These data demonstrate a physiologically important role for endogenous ET-1 and ET(A) receptors in the long-term regulation of arterial pressure and plasma ET-1 levels in the conscious nonhuman primate.

Selective antagonism of the ETA receptor, but not the ETB receptor, is protective against ischemic acute renal failure in rats.

We investigated the effects of ABT-627, a selective ETA-receptor antagonist, and A-192621, a selective ETB-receptor antagonist, on ischemic acute renal failure (ARF) in rats. Ischemic ARF was induced by clamping the left renal artery and vein for 45 min, 2 weeks after the contralateral nephrectomy. Renal function in untreated ARF rats markedly decreased at 24 h after reperfusion and thereafter tended to recover gradually. ABT-627 (1 mg/kg, i.v.) administration before ischemia markedly attenuated the renal dysfunction induced by the ischemia/reperfusion, whereas A-192621 (3 mg/kg, i.v.) pretreatment was without effect. Histopathological examination of the kidney of untreated ARF rats revealed severe renal damage such as tubular necrosis, proteinaceous casts in tubuli and medullary congestion. Histologically evident damage was improved by pretreatment with ABT-627, but not with A-192621. Daily oral administration of ABT-627 (10 mg/kg per day), but not A-192621 (30 mg/kg per day), given after the ischemia/reperfusion period also exerted protective effects. These findings clearly indicate that endothelin, acting via the ETA receptor, participates in the pathogenesis of ischemic ARF. Thus, selective ETA-receptor antagonism may be useful in the treatment of human ischemic ARF, whereas selective blockade of the ETB receptor will probably be ineffective.

Plasma endothelin-1 levels neither increase nor correlate with neurological scores, stroke risk factors, or outcome in patients with ischemic stroke.

BACKGROUND AND PURPOSE: Endothelins (ETs) are potent vasoconstrictors and may play a role in the pathophysiology of several diseases. Limited and controversial data exist on their role in human ischemic stroke. We planned a prospective, observational, and longitudinal clinical study to test whether ET-1 levels increase in various phases of ischemic stroke and whether the ET-1 levels correlate with neurological scores, stroke etiology, stroke risk factors, or final outcome. METHODS: We measured plasma ET-1 levels with a sandwich-enzyme immunoassay method in 101 consecutive patients with ischemic stroke on admission and 1 week, 1 month, and 3 months after stroke and in 101 sex- and age-matched control subjects. At each sampling, the patients underwent a complete neurological evaluation. All stroke risk factors were recorded, an array of laboratory tests were performed, and the subtype of ischemic stroke was determined. The patients were contacted 3 years later for prognostic determination. RESULTS: ET-1 levels in patients (2.4+/-1.3 pg/mL on admission, 2.2+/-1.4 pg/mL at 1 week, 2.1+/-1.4 pg/mL at 1 month, and 2.1+/-1.2 pg/mL at 3 months) were not different from those of the control subjects (2.2+/-0.9 pg/mL) at any time point. No correlation was found between the ET-1 levels and stroke etiology, stroke risk factors, stroke recurrence risk, age, sex, or neurological scores, except that ET-1 levels correlated with the use of warfarin and with body mass index. CONCLUSIONS: Plasma ET-1 levels were normal in patients with ischemic stroke. Our findings cannot exclude a role of ETs in the pathophysiology of ischemic stroke because plasma levels might not accurately reflect intracerebral concentrations, but they also do not support the occurrence of a major plasma ET-1 level increase at any phase of stroke. Our patient population is the largest ever reported in whom ET-1 levels were measured, but it consisted of mild and moderately ill patients with stroke due to the study design, of which the aim was long-term observation, which excludes severely ill patients.

Direct determination of endothelin receptor antagonist levels in plasma using a scintillation proximity assay.

An assay using scintillation proximity bead technology has been developed suitable for the quantitation of endothelin (ET) receptor antagonists in preclinical and clinical samples of plasma. The assay measures the competitive inhibition of radiolabelled ET-1 binding to ET(A) receptor membranes bound to wheat germ agglutinin (WGA)-coated scintillation proximity assay (SPA) beads in the presence of plasma containing A-127722, a potent orally active, ET(A) selective ET antagonist. The assay requires as little as 50 microl plasma and no extraction procedure is needed. The SPA methodology eliminates the need for the separation of bound from free ligand. Using this method, A-127722 could be directly quantified in rat plasma with a detection limit of 1 ng/ml.

ABT-627, an endothelin ET(A) receptor-selective antagonist, attenuates tactile allodynia in a diabetic rat model of neuropathic pain.

Tactile allodynia, the enhanced perception of pain in response to normally non-painful stimulation, represents a common complication of diabetic neuropathy. The activation of endothelin ET(A) receptors has been implicated in diabetes-induced reductions in peripheral neurovascularization and concomitant endoneurial hypoxia. Endothelin receptor activation has also been shown to alter the peripheral and central processing of nociceptive information. The present study was conducted to evaluate the antinociceptive effects of the novel endothelin ET(A) receptor-selective antagonist, 2R-(4-methoxyphenyl)-4S-(1,3-benzodioxol-5-yl)-1-(N, N-di(n-butyl)aminocarbonyl-methyl)-pyrrolidine-3R-carboxylic acid (ABT-627), in the streptozotocin-induced diabetic rat model of neuropathic pain. Rats were injected with 75 mg/kg streptozotocin (i. p.), and drug effects were assessed 8-12 weeks following streptozotocin treatment to allow for stabilization of blood glucose levels (>/=240 mg/dl) and tactile allodynia thresholds (

Pyrrolidine-3-carboxylic acids as endothelin antagonists. 4. Side chain conformational restriction leads to ET(B) selectivity.

When the dialkylacetamide side chain of the ET(A)-selective antagonist ABT-627 is replaced with a 2,6-dialkylacetanilide, the resultant analogues show a complete reversal of receptor selectivity, preferring ET(B) over ET(A). By optimizing the aniline substitution pattern, as well as the alkoxy group on the 2-aryl substituent, it is possible to prepare antagonists with subnanomolar affinity for ET(B) and with selectivities in excess of 4000-fold. A number of these compounds also show promising pharmacokinetic profiles; a useful balance of properties is found in A-192621 (38). Pharmacology studies with A-192621 serve to reveal the role of the ET(B) receptor in modulating blood pressure; the observed hypertensive response to persistent ET(B) blockade is consistent with previous postulates and indicates that ET(B)-selective antagonists may not be suitable as agents for long-term systemic therapy.

Design, synthesis, and activity of a series of pyrrolidine-3-carboxylic acid-based, highly specific, orally active ET(B) antagonists containing a diphenylmethylamine acetamide side chain.

The endothelin (ET)-B receptor subtype is expressed on vascular endothelial and smooth muscle cells and mediates both vasodilation and vasoconstriction. On the basis of the pharmacophore of the previously reported ET(A)-specific antagonist 1, (ABT-627), we are reporting the discovery of a novel series of highly specific, orally active ET(B) receptor antagonists. Replacing the dibutylaminoacetamide group of 1 with a diphenylmethylaminoacetamide group resulted in antagonist 2 with a complete reversal of receptor specificity. Structure-activity relationship studies revealed that ortho-alkylation of the phenyl rings could further increase ET(B) affinity and also boost the ET(A)/ET(B) activity ratio of the resulting antagonists. A similar antagonism selectivity profile could also be achieved when one of the phenyl rings of the acetamide side chain was replaced with an alkyl group, preferably a tert-butyl group (10h). Combining these features with modification of the 2-aryl group of the pyrrolidine core, we have identified a potent antagonist (9k, A-308165) with over 27 000-fold selectivity favoring the ET(B) receptor and an acceptable pharmacokinetic profile (F = 24%) in rats.

Discovery of a series of pyrrolidine-based endothelin receptor antagonists with enhanced ET(A) receptor selectivity.

Endothelins, ET-1, ET-2, and ET-3 are potent vasoconstricting and mitogenic 21-amino acid bicyclic peptides, which exert their effects upon binding to the ET(A) and ET(B) receptors. The ET(A) receptor mediates vasoconstriction and smooth muscle cell proliferation, and the ET(B) receptor mediates different effects in different tissues, including nitric oxide release from endothelial cells, and vasoconstriction in certain vascular cell types. Selective antagonists of endothelin receptor subtypes may prove useful in determining the role of endothelin in various tissue types and disease states, and hence as therapeutic agents for such diseases. The pyrrolidine carboxylic acid A-127722 has been disclosed as a potent and ET(A)-selective antagonist, and is currently undergoing clinical trials. In our efforts to find antagonists with altered selectivity (ET(A)-selective, ET(B)-selective, or nonselective), we investigated the SAR of the 2-substituent on the pyrrolidine. Compounds with alkyl groups at the 2-position possessed ET(A) selectivity improved over A-127722 (1400-fold selective), with the best of these compounds showing nearly 19,000-fold selectivity.

New bone formation in an osteoblastic tumor model is increased by endothelin-1 overexpression and decreased by endothelin A receptor blockade.

OBJECTIVES: The osteoblastic response of bone to metastatic prostate cancer is both characteristic and enigmatic. The potent vasoconstrictor endothelin-1 (ET-1), produced by prostate cancer, has been identified as a potential factor in new bone formation. METHODS: Using a novel method to quantitate new bone formation induced by the WISH tumor, we examined the effects of ET-1 overexpression and endothelin receptor antagonists on the osteoblastic response. RESULTS: WISH, a human tumor cell line derived from amnion, produces ET-1 mRNA and protein and induces abundant new bone formation and splenomegaly in vivo. Stable transfection of WISH with an ET-1 overexpression cDNA construct produced clones that secreted 18-fold more bioactive ET-1 than vector-only controls. After 14 days of growth in the lower leg of nu/nu mice, ET-1 overexpressing tumors produced significantly more new bone than vector-only controls. Conversely, areas of new bone formation were significantly less in animals treated with a selective endothelin A (ET(A)) receptor antagonist A127722. CONCLUSIONS: The activity of ET-1 in this osteoblastic model provides a unique target for therapy.

Perioperative plasma endothelin-1 and Big endothelin-1 concentrations in elderly patients undergoing major surgical procedures.

UNLABELLED: Plasma concentrations of the vasoconstrictor endothelin-1 (ET-1) increase during acute physiologic stress, but the role of ET-1 in the pathophysiology of stress remains largely undefined. Whether ET-1 mediates thermoregulatory changes in vasomotor tone is unknown. ET-1 and its more stable precursor, Big ET-1, were measured in plasma obtained at several perioperative time points from 95 consecutive elderly patients (mean age 70 +/- 1 yr) randomized to receive either normothermic or hypothermic perioperative care while undergoing major surgical procedures. In the postoperative period, there were no significant changes in plasma ET-1 concentrations, but Big ET-1 concentrations increased considerably (P < 0.0001). There were no significant differences in mean ET-1 or Big ET-1 levels in normothermic and hypothermic patients. Preoperative and postoperative ET-1 concentrations were significantly higher in patients with a history of hypertension (P < 0.002) and in those requiring treatment for postoperative hypertension (P < 0.003). Patients with cancer and those undergoing abdominal surgery had significantly higher Big ET-1 concentrations (P < 0.0001 and P < 0.003, respectively). These data support the hypothesis that Big ET-1 is a more sensitive measure of endothelin system activation after major surgery. Premorbid conditions and location and type of surgery influence perioperative ET-1/Big ET-1 concentrations. IMPLICATIONS: The endothelin response seems to be significantly associated with perioperative hemodynamic aberrations. The endothelin-1 (ET-1) precursor Big ET-1 is a more sensitive measure of the endothelin system activation in response to surgical stress than ET-1 alone. Thermoregulatory vasoconstriction in response to mild perioperative hypothermia occurs independently of the endothelin system.

Pharmacology of A-216546: a highly selective antagonist for endothelin ET(A) receptor.

Endothelins, 21-amino acid peptides involved in the pathogenesis of various diseases, bind to endothelin ET(A) and ET(B) receptors to initiate their effects. Here, we characterize the pharmacology of A-216546 ([2S-(2,2-dimethylpentyl)-4S-(7-methoxy-1,3-benzodioxol-5-yl )-1-(N,N-di(n-butyl) aminocarbonylmethyl)-pyrrolidine-3R-carboxylic acid), a potent antagonist with > 25,000-fold selectivity for the endothelin ET(A) receptor. A-216546 inhibited [125I]endothelin-1 binding to cloned human endothelin ET(A) and ET(B) receptors competitively with Ki of 0.46 and 13,000 nM, and blocked endothelin-1-induced arachidonic acid release and phosphatidylinositol hydrolysis with IC50 of 0.59 and 3 nM, respectively. In isolated vessels, A-216546 inhibited endothelin ET(A) receptor-mediated endothelin-1-induced vasoconstriction, and endothelin ET(B) receptor-mediated sarafotoxin 6c-induced vasoconstriction with pA2 of 8.29 and 4.57, respectively. A-216546 was orally available in rat, dog and monkey. In vivo, A-216546 dose-dependently blocked endothelin-1-induced pressor response in conscious rats. Maximal inhibition remained constant for at least 8 h after dosing. In conclusion, A-216546 is a potent, highly endothelin ET(A) receptor-selective and orally available antagonist, and will be useful for treating endothelin-1-mediated diseases.

Different contributions of endothelin-A and endothelin-B receptors in the pathogenesis of deoxycorticosterone acetate-salt-induced hypertension in rats.

We investigated the involvement of actions mediated by endothelin-A (ETA) and endothelin-B (ETB) receptors in the pathogenesis of deoxycorticosterone acetate (DOCA)-salt-induced hypertension in rats. Two weeks after the start of DOCA-salt treatment, rats were given ABT-627 (10 [mg/kg]/d), a selective ETA receptor antagonist; A-192621 (30 [mg/kg]/d), a selective ETB receptor antagonist; or their vehicle for 2 weeks. Uninephrectomized rats without DOCA-salt treatment served as controls. Treatment with DOCA and salt for 2 weeks led to a mild but significant hypertension; in vehicle-treated DOCA-salt rats, systolic blood pressure increased markedly after 3 to 4 weeks. Daily administration of ABT-627 for 2 weeks almost abolished any further increases in blood pressure, whereas A-192621 did not affect the development of DOCA-salt-induced hypertension. When the degree of vascular hypertrophy of the aorta was histochemically evaluated at 4 weeks, there were significant increases in wall thickness, wall area, and wall-to-lumen ratio in vehicle-treated DOCA-salt rats compared with uninephrectomized control rats. The development of vascular hypertrophy was markedly suppressed by ABT-627. In contrast, treatment with A-192621 significantly exaggerated these vascular changes. In vehicle-treated DOCA-salt rats, renal blood flow and creatinine clearance decreased, and urinary excretion of protein, blood urea nitrogen, fractional excretion of sodium, and urinary N-acetyl-beta-glucosaminidase activity increased. Such damage was overcome by treatment with ABT-627 but not with A-192621; indeed, the latter agent led to worsening of the renal dysfunction. Histopathologic examination of the kidney in vehicle-treated DOCA-salt rats revealed tubular dilatation and atrophy as well as thickening of small arteries. Such damage was reduced in animals given ABT-627, whereas more severe histopathologic changes were observed in A-192621-treated animals. These results strongly support the view that ETA receptor-mediated action plays an important role in the pathogenesis of DOCA-salt-induced hypertension. On the other hand, it seems likely that the ETB receptor-mediated action protects against vascular and renal injuries in this model of hypertension. A selective ETA receptor antagonist is likely to be useful for treatment of subjects with mineralocorticoid-dependent hypertension, whereas ETB-selective antagonism alone is detrimental to such cases.

The effects of diminishing albumin binding to some Endothelin receptor antagonists.

As a pharmacological class, Endothelin-A receptor (ET(A)) antagonists are highly bound (>98%) to serum albumin. In the presence of physiological concentrations of albumin, their affinities for ET(A) decrease 10 to 100 fold. We have prepared ET(A) antagonists which exhibit lower degrees of binding to albumin, while maintaining potency and selectivity for the ET(A) receptor. The protein induced IC50 shift is reduced or eliminated in this new series of compounds. The compounds also display altered in vivo and pharmacokinetic profiles which may be consistent with their lower degree of protein binding.

Pyrrolidine-3-carboxylic acids as endothelin antagonists. 3. Discovery of a potent, 2-nonaryl, highly selective ETA antagonist (A-216546).

Previously we have reported the discovery of ABT-627 (1, A-147627, active enantiomer of A-127722), a 2,4-diaryl substituted pyrrolidine-3-carboxylic acid based endothelin receptor-A antagonist. This compound binds to the ETA receptor with an affinity (Ki) of 0. 034 nM and with a 2000-fold selectivity for the ETA receptor versus the ETB receptor. We have expanded our structure-activity studies in this series, in an attempt to further increase the ETA selectivity. When the p-anisyl group of 1 was replaced by an n-pentyl group, the resultant antagonist 3 exhibited substantially increased ETB/ETA activity ratio, but a decreased ETA affinity. Structure-activity studies revealed that substitution and geometry of this alkyl group, and substitution on the benzodioxolyl ring, are important in optimizing this series of highly ETA selective antagonists. In particular, the combination of a (E)-2,2-dimethyl-3-pentenyl group and a 7-methoxy-1,3-benzodioxol-5-yl group provided hydrophobic compound 10b with subnanomolar affinity for human ETA receptor subtype and with an ETB/ETA activity ratio of over 130000. Meanwhile, synthetic efforts en route to olefinic compounds led to the discovery that 2-pyridylethyl (9o) and 2-(2-oxopyrrolidinyl)ethyl (9u) replacement of the p-anisyl group of 1yielded very hydrophilic ETA antagonists with potency and selectivity equal to those of 10b. On the basis of overall superior affinity, high selectivity for the ETA receptor (Ki, 0.46 nM for ETA and 13000 nM for ETB), and good oral bioavailability (48% in rats), A-216546 (10a) was selected as a potential clinical backup for 1.