Salah D. Kivlighn

A new method of measuring renal function in conscious rats without the use of radioisotopes.

The purpose of the current experiment was to develop fast and accurate assays for measuring glomerular filtration rate (GFR) and effective renal plasma flow (ERPF). An enzymatic method was developed for the determination of inulin, and a colorimetric method was developed for determination of p-aminohippurate (PAH) in the plasma and urine of rats. These assays are easily automated and do not require the use of radioisotopes or corrosive chemicals. Glomerular filtration rate was measured by the clearance of inulin, and effective renal plasma flow was measured by the clearance of PAH. Blood pressure, heart rate, and renal function (urine volume, electrolytes, GFR, and ERPF) were measured in conscious rats for 1.5 h prior to drug treatment and for 3 h after treatment. Baseline renal function was compared to historical data. Acute changes in GFR and ERPF following administration of the vasoconstrictor peptide endothelin-1 (ET-1) were accurately measured with results similar to those obtained with older methodologies. These new methods offer many advantages over previously described methods by eliminating the use of radioisotopes and harsh chemicals. In addition, these methods can be used with an automated instrument with high accuracy and precision. Therefore, these new methods can be used to accurately determine GFR and ERPF and are sensitive enough to detect acute changes in GFR and ERPF in conscious animals.

Potent imidazole angiotensinII antagonists: acyl sulfonamides and acyl sulfamides as tetrazole replacements

Abstract Acyl sulfonamides and acyl sulfamides were synthesized and their in vitro and in vivo biological properties evaluated. AT1 binding affinities for these potent AII antagonists were similar to their tetrazole analogs. An enhancement in AT2 potencies was observed, particularly with acyl sulfonamides or sulfamides bearing hydrophobic substituents.

Imidazo[4,5-b]pyridine-based AT1 / AT2 angiotensin II receptor antagonists

Abstract The structure-activity relationships of 6-amido-imidazol [4,5-b]pyridine-based angiostensin II antagonists ( V demonstrate that high affinity for the AT1 and AT2 receptors is largely dependent upon the R1 and R4 substituents. Of this series, L-162,441 and L-162,620 exhibits subnanomolar (IC50) binding affinities to both AT1 and AT2 receptors and potent antihypertensive effects in animals upon oral administration.

A new class of balanced AT1/AT2 angiotensin II antagonists: quinazolinone AII antagonists with acylsulfonamide and sulfonylcarbamate acidic functionalities

Abstract The structure activity relationships of a series of 2-alkyl-6-(acylamino)-3-[((2′-acylaminosulfonyl)biphenyl-4-yl)methyl]quinazolin-4-(3H)-ones were studied in order to identify balanced angiotensin II antagonists capable of potent binding to both AT 1 and AT 2 angiotensin receptor subtypes. The optimization of the substitution pattern led to the discovery of a potent, balanced quinazolinone antagonist L-162,393 , which displayed long lasting blockade of angiotensin pressor response in rats, dogs and rhesus monkeys.

Quinazolinones 2: QSAR and in vivo characterization of AT1 selective AII antagonists

Abstract The structure activity relatoionship, linear regression analysis and in vivo evaluation of a series of substituted 2-butyl-3-[(2′-tetrazol-5-yl)biphen-4-yl)methyl]quinazolin-4(1H)-ones as antagonists of the AT1 receptor for angiotensin II is presented. L-159,093 (2-butyl-6-(N-isopropyl-N-methyl-carbamoyl)amino-3-[(2′-tetrazol-5-yl)biphen-4-yl)methyl]quin azolin-4(1H)-one (IC50=0.1nM rabbit aorta) is shown to be a potent orally active AII antagonist in rats and rhesus.

The SAR of 6-(N-alkyl-N-acyl)-2-propyl-3-[(2′-tetrazol-5-yl)biphen-4-yl)methyl]-quinazolinones as balanced affinity antagonists of the human AT1 and AT2 receptors

Abstract Modification of the 6-N-alkyl-N-acyl groups of L-159,689, 6 6-(N-benzoyl-N-pentyl)-amino-2-propyl-3-[(2′-(tetrazol-5-yl)biphen-4-yl)methyl]quinazolin-4-(3H)one led to the identification of the 6-(N-benzoyl-N-(3-pyridylmethyl)) analog (L-162,537). L-162,537 had improved aqueous solubility and oral bioavailability in the dog. The SAR of this class of AT1 and AT2 ligands is discussed.

In vitro pharmacology of an angiotensin AT1 receptor antagonist with balanced affinity for AT2 receptors

L-163,017 (6-[benzoylamino]-7-methyl-2-propyl-3-[[2-(N-(3-methyl-1-butoxy) carbonylaminosulfonyl) [1,1]-biphenyl-4-yl]methyl]-3H-imidazo[4,5-b]pyridine) inhibited specific 125I-[Sar1, Ile8]angiotensin II binding to angiotensin AT1 receptor (Ki = 0.11-0.20 nM) in rabbit aorta, rat adrenal and human angiotensin AT1 receptor in CHO (Chinese hamster ovary transformed) cells and to AT2 receptor (Ki = 0.14-0.23 nM) in rat adrenal and brain receptors. L-163,017 also had a high affinity in the presence of bovine serum albumin (2 mg/ml), for angiotensin AT1 and AT2 receptors on human adrenal (Ki 3.9 and 4.3 nM), aorta (Ki 0.45 and 0.96 nM) and kidney (Ki 3.6 and 2.3 nM). The much higher Ki values in human tissues were likely due to the presence of bovine serum albumin in the binding assay buffer since L-163,017 had Ki values of 0.13 +/- 0.04 and 2.0 +/- 0.04 nM in the absence and presence of bovine serum albumin, respectively, in inhibiting 125I-[Sar1,Ile8]angiotensin II binding to angiotensin AT1 receptor in rat adrenal membranes. Scatchard analysis of 125I-[Sar1,Ile8]angiotensin II binding in the presence of bovine serum albumin (2 mg/ml) in rabbit aorta and bovine cerebellum indicated a competitive interaction of L-163,017 with angiotensin AT1 and AT2 receptors (Ki values 2.5 and 2.1 nM respectively). L-163,017 inhibited angiotensin II-induced aldosterone release in rat adrenal demonstrating that L-163,017 acted as a competitive antagonist (pA2 = 9.9) and lacked agonist activity. L-163,017 also inhibited angiotensin II responses in rat vascular tissues. The specificity of L-163,017 was shown by its lack of activity on the above functional responses produced by other agonists and in several binding assays.

Quinazolinone biphenyl acylsulfonamides: a potent new class of angiotensin-II receptor antagonists

Abstract A new series of quinazolinone-based AT 1 selective antagonists, bearing acylsulfonamides (-SO 2 NHCOR) as the tetrazole bioisosteres, was evaluated. While AT 1 potencies remained similar to the tetrazole analogs, the AT 2 receptor affinities were significantly improved with the introduction of acylsulfonamide groups. Several of these antagonists displayed improved in vivo properties.

Development of angiotensin II antagonists with equipotent affinity for human AT1 and AT2 receptor subtypes.

Abstract The quinazoline sulfonylcarbamate L-163,579 (9) is a potent, balanced antagonist of the binding of angiotensin II (Ang II) to human AT 1 and AT 2 receptors. This antagonist produces a long-lasting blockade of Ang II-induced pressor response in both rats and dogs after oral administration.

In vivo pharmacology of a novel AT1 selective angiotensin II receptor antagonist, MK-996

MK-996, N-(4-(5,7-dimethyl-2-ethyl-3H-imidazo[4,5-b]pyridin-3-yl- methyl)1,1-biphenyl-2-yl)-sulfonylbenzamide, is a potent, orally active, highly selective, nonpeptide angiotensin II (AII) receptor antagonist. MK-996 prevents the pressor response to intravenous AII in the conscious rat, dog, and rhesus monkey (ED50, mg/kg; oral/intravenous = 0.067/0.014, 0.035/0.017, and 0.1/0.036, respectively). In the anesthetized chimpanzee, MK-996 (1 mg/kg, iv) produces 100% (peak) inhibition of the AII pressor response and is still active (52%) at 24 h. To our knowledge this pharmacologic profile in the rat, dog, rhesus monkey, and chimpanzee presents the least species variability of any AII receptor antagonist yet described. Responses to methoxamine and arginine vasopressin are not affected by MK-996. In aortic coarcted (high renin) rats, MK-996 (3 mg/kg, by mouth) reduces blood pressure to normotensive (< 120 mm Hg) levels without reflex tachycardia. This dose of MK-996 reduces blood pressure to approximately the same level as both losartan (3 mg/kg, by mouth) and enalapril (3 mg/kg, by mouth) in this model. The duration of antihypertensive activity of MK-996 is similar to enalapril and shorter than losartan at the doses tested. Additionally, in the rat MK-996 does not potentiate the vasodepressor response to bradykinin and completely prevents the ability of AII to stimulate an increase in plasma levels of aldosterone. Therefore, MK-996 is a potent, orally active, nonpeptide AII receptor antagonist with a long duration of action, little species variability, and anti-hypertensive activity.