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A series of 4-hydroxy-3-methylsulfonanilido-1,2-diarylethylamines were prepared and evaluated for their human beta(3) adrenergic receptor agonist activity. SAR studies led to the identification of BMS-196085 (25), a potent beta(3) full agonist (K(i)=21 nM, 95% activation) with partial agonist (45%) activity at the beta(1) receptor. Based on its desirable in vitro and in vivo properties, BMS-196085 was chosen for clinical evaluation.

In a previous study, the depressor activity of combined selective inhibitors of neutral endopeptidase EC 3.4.24.11 (NEP) and angiotensin-converting enzyme (ACE) depended on the level of ACE inhibition, whereas the renal responses were determined by NEP inhibition. Our study confirmed that a mixed NEP/ACE inhibitor BMS-182657 ([S-(R*,R*)]-2,3,4,5-tetrahydro-3-[(2-mercapto-1-oxo-3- phenylpropyl)amino]-2-oxo-1H-benzazepine-1-acetic acid) reduced mean arterial pressure (MAP) when renin release was reduced by a sodium load, suggesting that the depressor response did not require suppression of endogenous angiotensin II generation. Furthermore, a pressor dose of 30 ng/min of angiotensin II was required to block the depressor response to BMS-182657 in the presence or absence of exogenous human atrial natriuretic peptide (hANP 99-126). Thirty ng/min of angiotensin II also significantly enhanced the natriuresis induced by hANP 99-126 after BMS-182657 administration. In contrast, a nonpressor dose of angiotensin II (3 ng/min) reduced basal sodium excretion and the natriuretic responses to exogenous hANP 99-126 in the presence or absence of BMS-182657. The potentiation of the urinary ANP and cyclic guanosine monophosphate (cGMP) responses to hANP 99-126 by BMS-182657 was similar for all doses of angiotensin II; therefore angiotensin did not alter the effects of BMS-182657 on ANP metabolism or cGMP accumulation in the kidney. In summary, the renal responses to mixed metalloprotease inhibitors were apparently mediated by ANP potentiation and were modulated by angiotensin II. The depressor activity depended on ACE inhibition but was not mediated solely by reductions in endogenous angiotensin II levels.

1. Inhibitors of neutral endopeptidase (NEP) EC 3.4.24.11 were developed to regulate endogenous levels of the natriuretic and vasodilatory hormone atrial natriuretic peptide (ANP). The selective NEP inhibitor SQ 28603 enhanced the increases in plasma ANP and urinary excretion of ANP, cyclic GMP and sodium stimulated by infusion of human ANP in conscious monkeys. SQ 28603 also potentiated the renal and depressor responses to rat brain natriuretic peptide (BNP) in conscious spontaneously hypertensive rats (SHR) and human BNP in conscious monkeys. Therefore, selective NEP inhibitors protected both natriuretic peptides from degradation in vivo and enhanced their biological activities. 2. Selective NEP inhibitors lowered blood pressure in conscious DOCA/salt hypertensive rats and SHR with antihypertensive activity similar to that of exogenous ANP. Furthermore, simultaneous treatment with an angiotensin converting enzyme (ACE) inhibitor enhanced the depressor activity of the NEP inhibitor in SHR. 3. SQ 28603 stimulated urinary excretion of cyclic GMP and sodium in a dose-related manner in conscious dogs with tachycardia-induced heart failure. Addition of the ACE inhibitor captopril significantly reduced blood pressure and systemic vascular resistance while sustaining sodium excretion and increasing cardiac output, glomerular filtration rate and renal blood flow. Therefore, combined NEP and ACE inhibition produced a unique haemodynamic and renal profile in dogs with pacing-induced heart failure. 4. The novel dual metalloprotease inhibitor BMS-182657 potentiated the renal responses to exogenous ANP and suppressed the pressor response to angiotensin I in conscious monkeys, indicating in vivo inhibition of both NEP and ACE.(ABSTRACT TRUNCATED AT 250 WORDS)

1. The effects of sodium supplements on the renal responses to human atrial natriuretic peptide (hANP 99-126) and to the selective inhibitors of neutral endopeptidase 3.4.24.11 (NEP) SQ 28,603 and candoxatrilat were determined in conscious monkeys. 2. When the monkeys' diet was changed from 0.55% sodium to 1.1% sodium, the natriuretic response to 100 mumol/kg intravenous of SQ 28,603 increased from 665 +/- 64 to 1015 +/- 224 mu Eq/3 h. An acute oral load of 25 mEq sodium significantly increased the natriuresis stimulated by 300 mumol/kg, p.o., of SQ 28,603 from 700 +/- 332 mu Eq/3 h in normal monkey to 2437 +/- 841 mu Eq/3 h. Therefore, the non-human primate model was appropriate for investigating the effects of sodium loads on the urinary ANP and cGMP responses to exogenous ANP in the presence and absence of NEP inhibitors. 3. Graded intravenous infusions of saline increased basal urine volume and excretion of sodium and ANP. Salt supplements enhanced the diuretic, natriuretic and ANP responses to 0.3 nmol/kg intravenous of hANP 99-126 in monkeys treated with vehicle or 10 mumol/kg intravenous of candoxatrilat. The sodium and ANP excretions stimulated by hANP 99-126 were positively correlated with each other and with the calculated intravenous sodium load in the presence or absence of candoxatrilat. 4. SQ 28,603 and candoxatrilat (0.3 to 10 mumol/kg intravenous) each produced significant, dose-related potentiation of the natriuretic, cGMP and ANP responses to 0.3 nmol/kg intravenous of hANP 99-126 in monkeys receiving 5 mL/kg+0.2 mL/min saline. In addition, the highest dose of SQ 28,603 produced significant depressor activity. 5. In conclusion, the increased natriuretic activity of hANP 99-126 in sodium loaded monkeys was mediated, in part, by increased ANP delivery to the guanylate cyclase linked ANP receptors in the distal renal tubules.

The potent neutral endopeptidase inhibitor SQ 28 603 (N-(2-(mercaptomethyl)-1-oxo-3-phenylpropyl)-β-alanine) significantly increased excretion of sodium from 4.9 ± 2.3 to 14.3 ± 2.1 μequiv./min and cyclic 3′,5′-guanosine monophosphate from 118 ± 13 to 179 ± 18 pmol/min after intravenous administration of 300 μmol/kg (~80 mg/kg) in conscious female cynomolgus monkeys. SQ 28 603 did not change blood pressure or plasma atrial natriuretic peptide concentrations in the normal monkeys. In contrast, 1-h infusions of 3, 10, or 30 pmol∙kg−1∙min−1 of human atrial natriuretic peptide lowered blood pressure by −3 ± 4, −9 ± 4, and −27 ± 3 mmHg (1 mmHg = 133.322 Pa), increased cyclic guanosine monophosphate excretion from 78 ± 11 to 90 ± 6, 216 ± 33, and 531 ± 41 pmol/min, and raised plasma atrial natriuretic peptide from 7.2 ± 0.7 to 21 ± 4, 62 ± 12, and 192 ± 35 fmol/mL without affecting sodium excretion. In monkeys receiving 10 pmol∙kg−1∙min−1 of atrial natriuretic peptide, 300 μmol/kg of SQ 28 603 reduced mean arterial pressure by −13 ± 5 mmHg and increased sodium excretion from 6.6 ± 3.2 to 31.3 ± 6.0 μequiv./min, cyclic guanosine monophosphate excretion from 342 ± 68 to 1144 ± 418 pmol/min, and plasma atrial natriuretic peptide from 124 ± 8 to 262 ± 52 fmol/mL. In conclusion, SQ 28 603 stimulated renal excretory function in conscious monkeys, presumably by preventing the degradation of atrial natriuretic peptide by neutral endopeptidase.Key words: atrial natriuretic peptide, neutral endopeptidase, natriuresis, cyclic guanosine monophosphate.

The specific neutral endopeptidase (NEP) inhibitor, SQ 29,072 (7-[2-(mercaptomethyl)-1-oxo-3-phenylpropyl]amino]heptanoic acid), was studied in conscious spontaneously hypertensive rats (SHRs) and in DOCA/salt hypertensive rats during inhibition of angiotensin-converting enzyme (ACE) activity with captopril or SQ 27,519 (the free acid of fosinopril). In the SHR, the maximal depressor responses to the combination of SQ 29,072 and SQ 27,519 (-44 +/- 4 mm Hg) were greater than the responses to any of the inhibitors given alone (-26 +/- 5, -40 +/- 10, and -28 +/- 6 mm Hg for SQ 29,072, captopril, and SQ 27,519, respectively). In contrast, the maximal antihypertensive activities of SQ 29,072 were the same in conscious DOCA/salt hypertensive rats infused with saline, captopril, or SQ 27,519 (-54 +/- 10, -51 +/- 8, and -58 +/- 11 mm Hg, respectively), indicating a lack of synergism in this model. In agreement, SQ 28,133 [N-[2-(mercaptomethyl)-1-oxo-3-phenylpropyl]-L-leucine], a compound that inhibits both NEP and ACE, elicited significant depressor activities in both SHR and DOCA/salt hypertensive rats. In conclusion, a selective NEP inhibitor enhanced the depressor activity of ACE inhibitors in the conscious SHR, indicating that these agents may be effectively combined for treatment of some types of hypertension.

Efforts to further elucidate structure-activity relationships (SAR) within our previously disclosed series of beta-quaternary amino acid linked l-cis-4,5-methanoprolinenitrile dipeptidyl peptidase IV (DPP-IV) inhibitors led to the investigation of vinyl substitution at the beta-position of alpha-cycloalkyl-substituted glycines. Despite poor systemic exposure, vinyl-substituted compounds showed extended duration of action in acute rat ex vivo plasma DPP-IV inhibition models. Oxygenated putative metabolites were prepared and were shown to exhibit the potency and extended duration of action of their precursors in efficacy models measuring glucose clearance in Zucker(fa/fa) rats. Extension of this approach to adamantylglycine-derived inhibitors led to the discovery of highly potent inhibitors, including hydroxyadamantyl compound BMS-477118 (saxagliptin), a highly efficacious, stable, and long-acting DPP-IV inhibitor, which is currently undergoing clinical trials for treatment of type 2 diabetes.

The natriuretic and depressor responses to novel dual inhibitors of neutral endopeptidase (NEP) EC 3.4.24.11 and angiotensin-converting enzyme (ACE) were used to assess their activity in conscious cynomolgus monkeys. A survey of mercaptopropanoyl inhibitors revealed that compounds containing alanylproline or certain surrogates reduced blood pressure and increased sodium excretion, indicating a desirable profile of in vivo activity. Additional compound evaluation required specific in vivo assays for NEP and ACE inhibition. Accordingly, the potency of novel inhibitors against NEP and ACE were determined in conscious monkeys by the potentiation of the natriuretic activity of exogenous human atrial natriuretic peptide and inhibition of the pressor response to angiotensin I, respectively. This strategy led to the discovery that optimal in vivo activity was achieved when the mercaptopropanoyl group was replaced with mercaptoacetyl and the C-terminal alanylproline was replaced with conformationally constrained dipeptidomimetics. This work culminated in the identification of BMS-182657 as a prototypic dual NEP/ACE inhibitor with a highly desirable profile of in vivo pharmacology.

The depressor and renal responses to C-type natriuretic peptide (CNP) were determined in conscious cynomolgus monkeys treated with vehicle or inhibitors of neutral endopeptidase EC 3.4.24.11 (NEP) and angiotensin-converting enzyme (ACE). The NEP inhibitor SQ 28603 (100 mumol/kg intravenously, i.v.) significantly (p < 0.05) enhanced the depressor responses to 1 and 10 nmol/ kg i.v. CNP from -2 +/- 3 to -22 +/- 10 mm Hg and from -16 +/- 4 to -66 +/- 4 mm Hg, respectively. SQ 28603 also significantly increased the cyclic GMP responses to 1 and 10 nmol/kg CNP from 1.4 +/- 1.6 to 11.0 +/- 2.0 nmol/2 h and from 4.2 +/- 0.5 to 53.3 +/- 12.1 nmol/2 h, respectively. Furthermore, the NEP inhibitor significantly increased the natriuretic activity of 1 and 10 nmol/kg i.v. CNP from 235 +/- 99 to 760 +/- 60 microEq/2 h and from 399 +/- 208 to 1,036 +/- 79 microEq/2 h, respectively. A positive correlation between the cumulative natriuretic and cyclic GMP responses suggested a cyclic GMP-mediated mechanism. These data are consistent with the protection of CNP from degradation by renal NEP. Inhibition of ACE by 100 mumol/kg i.v. captopril did not significantly alter the depressor or renal activities of 1 nmol/kg of CNP, neither did it alter the potentiation of CNP activity by SQ 28603. The potentiation of the depressor, cyclic GMP, and natriuretic responses to CNP in nonhuman primates by SQ 28603 suggested that NEP is an important mechanism for in vivo inactivation of natriuretic peptides, including CNP.

1. There are at least two subtypes of vascular endothelin (ET) receptors. Stimulation of the ETA receptors on vascular smooth muscle cells leads to vasoconstriction, whereas activation of the ETB receptors on endothelial cells elicits vasodilatation. Several reports in the literature have suggested the presence of a vasoconstrictor non-ETA receptor on vascular smooth muscle which has pharmacological similarities to the ETB receptor. The present study was undertaken to determine the location of this ETB-like receptor within the vascular system. 2. Fourteen vascular smooth muscle preparations from six species were used to determine the effect of the ETA receptor antagonist, BQ-123, on concentration-response curves elicited by ET-1 and the ability of the ETB receptor agonist, sarafotoxin S6c, to cause contraction. The vessels fell into two categories. One group was sensitive to BQ-123 and insensitive to sarafotoxin S6c and, thus, probably contained ETA receptors. The other group, with vasoconstrictor ETB-like receptors, was insensitive to BQ-123 and sensitive to sarafotoxin S6c. 3. Vessels from cynomolgus monkeys, when studied in vitro, appeared to contain primarily ETA receptors, although the potency of BQ-123 was quite variable, suggesting the possibility of ETA receptor subtypes. In contrast, both ET-1 and sarafotoxin S6c, given as intravenous injections in conscious monkeys, produced transient, equipotent, and dose-related increases in blood pressure. The highest dose of sarafotoxin S6c (1 nmol kg-1, i.v.) also caused a marked secondary depressor response (-80 +/- 6 mmHg) that lasted approximately 10 min. The pressor responses suggest that the vasoconstrictor ETB-like receptors are present in cynomolgus monkeys. 4. The data suggest the presence of two distinct vasoconstrictor ET receptor subtypes on smooth muscle cells. The ETA receptors are primarily located on the high pressure side of the circulation. The vasoconstrictor ETB-like receptors appear to be concentrated on the low pressure side.