Pharmacological profile of BIBN4096BS, the first selective small molecule CGRP antagonist.

Calcitonin gene-related peptide (CGRP) is one of the most potent endogenous vasodilators known. This peptide is increased during migraine attacks and has been implicated in the pathogenesis of migraine headache. Here we report on the first small molecule selective CGRP antagonist: BIBN4096BS. In vitro, this compound is extremely potent at primate CGRP receptors exhibiting an affinity (Ki) for human CGRP receptors of 14.4 +/- 6.3 (n = 4) pM. In an in vivo model, BIBN4096BS in doses between 1 and 30 micrograms kg-1 (i.v.) inhibited the effects of CGRP, released by stimulation of the trigeminal ganglion, on facial blood flow in marmoset monkeys. It is concluded that BIBN4096BS is a potent and selective CGRP antagonist.

High throughput drug profiling.

High throughput screening has significantly contributed to advances in drug discovery. The great increase in the number of samples screened has been accompanied by increases in costs and in the data required for the investigated compounds. High throughput profiling addresses the issues of compound selectivity and specificity. It combines conventional screening with data mining technologies to give a full set of data, enabling development candidates to be more fully compared.

CGRP 27-37 analogues with high affinity to the CGRP1 receptor show antagonistic properties in a rat blood flow assay.

CGRP Y0-28-37 is known as a selective CGRP1 receptor antagonist. We succeeded in optimising the CGRP1 receptor affinity of this fragment by multiple amino acid replacement. The analogues [p34, F35]CGRP 27-37 and [D31, p34, F35]CGRP 27-37 exhibit a 100-fold increased affinity compared to the unmodified segment. Receptor binding studies were performed with human neuroblastoma cells SK-N-MC, which selectively express the hCGRP1 receptor. Blood flow, which is increased by exogenous CGRP, was measured in the right femoral artery. Preincubation of the rats with [p34, F35]CGRP 27-37 and [D31, p34, F35]CGRP 27-37 led to a significant decrease in CGRP induced increase in vascular conductance indicating the antagonistic properties of these compounds. Interestingly, an exchange of the amino acid Asn31 to Asp31 in [p34, F35]CGRP 27-37 shortened the period of the antagonistic effect significantly, suggestive of a different rate of metabolism for the two ligands. Secondary structure investigations obtained by circular dichroism measurements revealed that an increase in ordered structure correlates with high binding affinity.

From micromolar to nanomolar affinity: a systematic approach to identify the binding site of CGRP at the human calcitonin gene-related peptide 1 receptor.

CGRP Y0-28-37 is known as a selective CGRP1 receptor antagonist. In order to elucidate the essential requirements for its receptor interaction, we performed a variety of systematic approaches by modifying the C-terminal segments CGRP Y0-28-37 and CGRP 27-37. N-Terminal and C-terminal segments have been synthesized, as well as chimeras which combine segments of CGRP, adrenomedullin, and amylin. Furthermore, we carried out an Ala scan, a Phe scan, a D-amino acid scan and a Pro scan of CGRP 27-37. Additionally, single amino acids were replaced by those with similar biophysical properties. Receptor binding studies of all analogs were performed at human neuroblastoma cells SK-N-MC, which selectively express the hCGRP1 receptor. On the basis of the obtained results, we synthesized a series of ligands with multiple amino acid replacements in order to optimize the exchange at each position. This approach yielded to a series of high affinity ligands, including [D31,P34,F35] CGRP 27-37 which exhibits a 100-fold increased affinity compared to the unmodified segment. So far, this is the smallest CGRP analog that shows affinity in the nanomolar range.

BIBP 3226, the first selective neuropeptide Y1 receptor antagonist: a review of its pharmacological properties.

Based on the assumption that the pharmacophoric groups interacting with the Y1 receptor are located in the C-terminal part of neuropeptide Y, low molecular weight compounds with high affinity and selectivity for the Y1 receptor were designed and synthesized. The prototype BIBP 3226 possesses affinity for the Y1 receptor in the nanomolar range. In addition, this compound is selective displaying rather low affinity for Y2, Y3, Y4 and a set of 60 other receptors. Both biochemical and pharmacological studies showed that BIBP 3226 behaves as a competitive antagonist. Using BIBP 3226 it was possible to investigate the role of NPY and/or Y1 receptors in blood pressure regulation. The interesting observation was that antagonism to Y1 receptors had no major influence on the basal blood pressure but attenuated stress induced hypertension. This strongly supports the hypothesis that NPY is mainly released during stress involving intense sympathetic nervous system activation. Moreover, BIBP 3226 can be used to characterize NPY receptor subtypes. For instance, we were able to show that presynaptic NPY receptors mediating catecholamine release do not solely belong to the Y2 subtype, but that presynaptic Y1 receptors also exist. In conclusion, BIBP 3226 has been shown to be an important tool for the elucidation of the physiological role of Y1 receptors in the cardiovascular system.

Pharmacological characterization of the selective nonpeptide neuropeptide Y Y1 receptor antagonist BIBP 3226.

The present study was undertaken to investigate the in vitro and in vivo pharmacological profile of the novel, nonpeptide neuropeptide Y (NPY) Y1-selective antagonist, BIBP 3226 [(R)-N2-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]-D-arginine-am ide], and a recently described peptidic structure [Ile-Glu-Pro-Orn-Tyr-Arg-Leu-Arg-Tyr-NH2, cyclic (2,4'), (2',4)-diamide]. BIBP 3226 antagonized the NPY Y1 receptor-mediated decrease in the twitch response in the rabbit vas deferens preparation with a pKb value of 6.98 +/- 0.06 (n = 16). It showed no affinity (EC50 > 1 microM) for NPY Y2 receptors in the rat vas deferens. NPY-induced increases in perfusion pressure in the isolated perfused rat kidney and rabbit ear preparations were antagonized with IC50 values of 26.8 +/- 4.5 (n = 4) and 214 +/- 30 nM (n = 4), respectively. The NPY-mediated potentiation of the noradrenaline elicited increase in perfusion pressure in the rat mesenteric bed was antagonized with an IC50 value of 976 (542-1760) nM. The NPY-induced increase in blood pressure in the pithed rat was inhibited by BIBP 3226 dose-dependently (ED50 = 0.11 +/- 0.03 mg/kg i.v.), whereas no effect of BIBP 3226 (1 mg/kg i.v.) was observed for the noradrenaline-, angiotensin-, endothelin- or vasopressin-induced pressor response. The data presented demonstrate that BIBP 3226 is a competitive and NPY Y1-selective antagonist. The peptidic compound proved to possess high potency for NPY Y1 receptors, but showed both agonistic as well as antagonistic properties.(ABSTRACT TRUNCATED AT 250 WORDS)

Subtype selectivity and antagonistic profile of the nonpeptide Y1 receptor antagonist BIBP 3226.

In the present study, the subtype specificity and species selectivity of the nonpeptide BIBP 3226, as well as its in vitro antagonism of neuropeptide Y (NPY)-mediated second messengers have been investigated. Radiolabeled NPY is potently displaced by BIBP 3226 [(R)-N2-(diphenylacetyl)-N-[(4-hydroxyphenylmethyl]-D- arginine amide] on human Y1 receptor expressing Chinese hamster ovary-K1 cells (Ki = 0.47 +/- 0.07 nM). SK-N-MC human neuroblastoma cells (Ki = 5.1 +/- 0.5 nM) and the rat parietal cortex membranes (Ki = 6.8 +/- 0.7 nM). The interaction of BIBP 3226 with the Y1 receptor is stereoselective, because the (S)-enantiomer of the (R)-configured BIBP 3226 displays almost no affinity (Ki > 10,000 nM). In contrast, concentrations up to 10 microM BIBP 3226 do not displace [125I]NPY from the human Y2 receptor (neuroblastoma cell line SMS-KAN), the rabbit Y2 receptor (kidney) and the rat Y2 receptor (hippocampus). Functional antagonism could be shown for the human Y1 receptor: 0.1 microM BIBP 3226 antagonizes the NPY induced Ca++ mobilization (pKb = 7.5 +/- 0.17) as well as the NPY-mediated inhibition of cyclic AMP synthesis (pKb = 8.2 +/- 0.24) in SK-N-MC cells. In contrast, none of the formerly described putative antagonists PYX-2, [D-Trp32]NPY and benextramine could be characterized as high affinity Y1 receptor antagonists. The 18 amino acid NPY analog EXBP 68 Ile-Glu-Pro-Orn-Tyr-Arg-Leu-Arg-Tyr-NH2, cyclic (2,4'), (2',4')-diamide] displayed Y1-selective affinity with in vitro antagonistic properties (Ki = 0.33 +/- 0.04 nM and pKb = 8.4 +/- 0.07) in SK-N-MC cells. Therefore, BIBP 3226 is the first potent and subtype-selective nonpeptide Y1 receptor antagonist.

Labeling of neuropeptide Y receptors in SK-N-MC cells using the novel, nonpeptide Y1 receptor-selective antagonist [3H]BIBP3226.

The binding of tritium-labelled BIBP3226, N2-(diphenylacetyl)-N-[(4-hydroxy-phenyl)methyl]-D-arginine amide, to human neuroblastoma SK-N-MC cells was investigated. [3H]BIBP3226 reversibly binds to neuropeptide Y receptors of the Y1 subtype expressed in SK-N-MC cells with a KD of 2.1 +/- 0.3 nM (mean +/- S.E.M., n = 3) and a Bmax of 58,400 +/- 1100 sites/cell. Non-specific binding did not exceed 30% of the total radioactivity bound at KD. In competition experiments [3H]BIBP3226 is concentration-dependently displaced by neuropeptide Y and its peptide analogues with an affinity pattern neuropeptide Y = [Leu31, Pro34]neuropeptide Y >> neuropeptide Y-(18-36). This rank order of potencies is consistent with the interaction of [3H]BIBP3226 with neuropeptide Y receptors of the Y1 subtype. Therefore, [3H]BIBP3226 can be used as selective ligand to study neuropeptide Y Y1 receptors.

Effects of pimobendan and its metabolite on myofibrillar calcium responsiveness and ATPase activity in the presence of inorganic phosphate.

The effects of the cardiotonic agent pimobendan (CAS 118428-36-7, UD-CG 115 BS) and its main metabolite UD-CG 212 on dog cardiac myofibrillar calcium responsiveness and ATPase activity were studied at nominal free inorganic phosphate (Pi) and at 5 mmol/l Pi. A rightward shift of the pCa-tension relationship with a marked depression of maximal tension was observed in the presence of 5 mmol/l Pi. Pimobendan increased myofibrillar calcium responsiveness at concentrations > or = 10(-5) mol/l. These effects of pimobendan were significantly greater at 5 mmol/l Pi than at nominally free Pi. UD-CG 212 had no influence on myofibrillar calcium responsiveness at nominally free Pi, however, significant effects were observed at 10(-9) mol/l UD-CG 212 in the presence of 5 mmol/l Pi. UD-CG 212 (10(-8) mol/l) did not influence myofibrillar ATPase activity at pCa's 6.23, 5.99, and 4.36 with or without 5 mmol/l Pi, whereas pimobendan (10(-4) mol/l) had an effect only at pCa = 5.99 (without Pi) and pCa = 4.36 (+ 5 mmol/l Pi). The data suggest that the increase in myofibrillar calcium responsiveness at submaximal calcium concentrations by pimobendan and UD-CG 212 in the presence of 5 mmol/l Pi is brought about by a change in cross-bridge kinetics or by enhancement of thin filament activation by adjacent strong cross-bridges. At maximal calcium activation, pimobendan may additionally increase the population of strong cross-bridges.

Adrenomedullin mediates vasodilation via CGRP1 receptors.

Adrenomedullin is a recently discovered 52 amino acid polypeptide with potent hypotensive activity. The peptide possesses 21% homology with the amino acid sequence of human calcitonin gene-related peptide-alpha (hCGRP-alpha). In 125I-hCGRP-alpha receptor binding experiments using membranes from human neuroblastoma cells (SK-N-MC) adrenomedullin is a potent competitor with a Ki of 0.37 nM. In SK-N-MC cells hCGRP-alpha and adrenomedullin concentration-dependently increase cAMP levels with -logEC50 values of 9.65 and 7.75, respectively. Both responses were attenuated in the presence of 30 nM CGRP[8-37], a CGRP1 receptor antagonist. In isolated rat hearts, perfused at constant flow, bolus infusion of adrenomedullin (1 to 100 nM) resulted in a concentration-dependent, pronounced and long-lasting vasodilation with an approximate EC50 of about 3 nM. This effect was markedly attenuated in the presence of 100 nM CGRP[8-37]. In this model, bolus infusion of hCGRP-alpha (0.01 to 100 nM) evoked a comparable vasodilation with an approximate EC50 of 0.5 nM. This effect was also potently inhibited in the presence of CGRP[8-37]. These results suggest that adrenomedullin-mediated vasodilation is linked to the activation of CGRP1 receptors in the coronary vascular system.

The first highly potent and selective non-peptide neuropeptide Y Y1 receptor antagonist: BIBP3226.

The design and subsequent in vitro and in vivo biological characterisation of the first potent and selective non-peptide neuropeptide Y Y1 receptor antagonist, BIBP3226 ((R)-N2-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]-argininami de) is reported. BIBP3226 displaced 125I-labelled neuropeptide Y with high affinity (Ki = 7 nM) from the human neuropeptide Y Y1 receptor and proved to be highly selective. BIBP3226 displayed potent antagonistic properties both in in vitro and in vivo models and thus represents the first selective non-peptide neuropeptide Y Y1 receptor antagonist.

Mutations in transmembrane segment VII of the AT1 receptor differentiate between closely related insurmountable and competitive angiotensin antagonists.

Chimeric constructs between the human and the Xenopus laevis AT1 receptor have demonstrated, that the binding of non-peptide angiotensin antagonists is dependent on non-conserved residues located deep in transmembrane segment VII of the AT1 receptor. Here we have studied four pairs of closely related antagonists each consisting of a competitive and an insurmountable compound differentiated by one out of three different types of minor chemical modifications. None of the antagonists bound to the Xenopus receptor and the binding of all of the compounds to the human receptor was severely impaired by the introduction of non-conserved residues from transmembrane segment VII of the Xenopus receptor. In all four pairs of antagonists the competitive compound was affected more by these substitutions than the corresponding insurmountable one (209 vs. 22, 281 vs. 29, 290 vs. 29 and 992 vs. 325-fold increase in Ki values). A similar pattern was observed in response to substitution of a single non-conserved residue in transmembrane segment VII, Asn295 to Ser. These results indicate that a common molecular mechanism distinguishes the interaction of insurmountable and competitive antagonists with the AT1 receptor.

Pharmacological profile of selective muscarinic receptor antagonists on guinea-pig ileal smooth muscle.

The present study examined the effects of a series of tricyclic muscarinic receptor antagonists on muscarinic receptors present in the guinea-pig ileum, both in vitro and in vivo. The selectivity profiles of these antagonists and that of atropine were determined by their affinity for cortical muscarinic M1, cardiac M2 and submandibular M3 receptors and for m4 receptors expressed in CHO cells. The compounds pirenzepine, UH-AH 37, AQ-RA 391 and AQ-RA 618 possessed high affinity (pKi 7.94-8.22) for muscarinic M1 receptors. Pirenzepine exhibited the most pronounced muscarinic M1 selectivity. AF-DX 384 and AQ-RA 741 possessed an approximately 10-fold higher affinity for the cardiac muscarinic M2 receptor than AF-DX 116. However, both compounds also exhibited high affinity for muscarinic m4 receptors. High affinity for muscarinic M3 and m4 receptors was observed for UH-AH 37, AQ-RA 391 and AQ-RA 681. The antagonists were then tested for their interaction with the muscarinic receptors which are responsible for the methacholine-induced contraction of longitudinal muscle in vitro, circular muscle in vivo and muscarinic receptors which mediate the distension-evoked ascending reflex contraction of circular muscle in vitro. Compounds showing high affinity for muscarinic M3 receptors (e.g. AQ-RA 618) were the most potent antagonists in the functional experiments. Comparison of the binding displacement data with the functional results indicates that the effects of methacholine on the longitudinal and circular muscle of the guinea-pig ileum were predominantly mediated by muscarinic M3-type receptors. In contrast, the correlation between muscarinic M2 receptor affinity and antagonism of muscarinic receptors in the ileum was very weak.

Effects on binding characteristics and renal function of the novel, non-peptide angiotensin II antagonist BIBR277 in the rat.

OBJECTIVE: To characterize the interaction of BIBR277, a new angiotensin II (Ang II) type 1 receptor (AT1)-selective antagonist, with rat renal Ang II receptors and to investigate its effects on renal function in vitro and in vivo. METHODS: The binding characteristics of BIBR277 in the rat kidney were evaluated in [125I]-Ang II displacement and autoradiographic studies. Renal function was assessed in vitro in the isolated, constant-pressure perfused rat kidney and in vivo in anaesthetized rats. RESULTS: In rat kidney cortical membrane preparations BIBR277 binds to a single population of Ang II receptors, which are of the AT1 subtype. In autoradiographic studies specific [125I]-(Sar1,Ile8)-Ang II binding to the rat kidney glomeruli, renal cortex and medulla was completely inhibited by 1 mumol/l BIBR277. In isolated rat kidneys BIBR277 (0.01, 0.1 and 1 mumol/l) increased perfusate flow, urinary flow and glomerular filtration rate concentration-dependently by 115, 130 and 112% of the control value, respectively. This effect was not blocked in the presence of indomethacin (10 mumol/l). Frusemide (10 mumol/l) increased urinary flow in the isolated kidney to about 140% of the control value. The diuretic effect of frusemide was significantly increased to 180 and 200% of the control value in the presence of 0.1 and 1.0 mumol/l BIBR277, respectively. Captopril (10 mumol/l) had no effect on perfusate flow, urinary flow or glomerular filtration rate, and did not increase frusemide-induced diuresis in this preparation. BIBR277 was also administered intravenously to anaesthetized rats at doses of 0.1, 0.3 and 1 mg/kg. BIBR277 had no effect on the heart rate, but decreased the blood pressure significantly at both higher doses. At the 0.3 mg/kg dose, the urinary flow and sodium excretion increased significantly (twofold and 2.47-fold, respectively) compared with the vehicle-treated group, but not increase in the urinary flow and sodium excretion was observed at the highest dose. Urinary potassium excretion was not significantly affected at all doses. CONCLUSIONS: These results show that BIBR277 potently interacts with rat renal AT1 receptors. BIBR277 shows diuretic effects both in vitro and in vivo. In anaesthetized rats BIBR277 also promotes sodium excretion without affecting potassium excretion.

Characterization of muscarinic receptors in guinea-pig uterus.

To characterize the muscarinic receptor present in guinea-pig uterus smooth muscle the affinities of a series of 27 muscarinic receptor antagonists for M1 (rat cortex), M2 (rat heart), M3 (rat submandibular gland), m4 (transfected in CHO cells) and muscarinic binding sites in guinea-pig uterus smooth muscle were determined in radioligand binding studies. In addition, functional experiments were performed to assess pKB values of the antagonist for muscarinic receptors in guinea-pig atrium and uterus. The results obtained are consistent with the presence of M2 receptors in the uterus through which the functional contractile response is mediated. Correlation coefficients of 0.98, 0.91 and 0.91 were calculated for the following linear regressions: pKi uterus vs. pKi M2, pKB uterus vs. pKi M2 and pKB uterus vs. pKB atrium. This study also revealed that the compounds dicyclomine, DAU 5884, DAU 6202 as well as AQ-RA 721 could distinguish m4 from M2 sites and are therefore important tools to characterize muscarinic receptor subtypes. In addition, DAU 5884 and DAU 6202 have been identified as highly potent M1 selective antagonists.

6-Substituted benzimidazoles as new nonpeptide angiotensin II receptor antagonists: synthesis, biological activity, and structure-activity relationships.

Starting from the recently reported nonpeptidic angiotensin II (AII) receptor antagonists DuP753 (1) and Exp 7711 (2), we have designed and investigated novel substituted benzimidazoles. Systemic variation of several substituents at the benzimidazole ring positions 4-7 led to the finding that substitution in position 6 with acylamino groups results in highly active AII antagonists. Compounds with 6-membered lactam or sultam substituents in position 6 of benzimidazole showed receptor activities in the low nanomolar range but were only weakly active when given orally to rats. In contrast, analogous substitution of the benzimidazole moiety with basic heterocycles resulted in potent AII antagonists which were also well absorbed after oral application. The most active compound of this series, 33 (BIBR 277), was selected as a candidate for clinical development. On the basis of molecular modeling studies a binding model of this new class of AII antagonists to the AT1 receptor is proposed.

Characterization of BIBN 99: a lipophilic and selective muscarinic M2 receptor antagonist.

The present study was designed to characterize the receptor selectivity profile of the novel muscarinic M2 receptor antagonist BIBN 99 (5,11-dihydro-8-chloro-11-[[4-[3-[(2,2-dimethyl-1- oxopentyl)ethylamino]propyl]-1-piperidinyl]acetyl]-6H- pyrido[2,3-b][1,4]benzodiazepin-6-one). In radioligand binding studies BIBN 99 showed high affinity for m2/M2 sites (pKi = 7.52/7.57), intermediate affinity for m4 sites (pKi = 6.76) and low affinity for m1/M1 (pKi = 5.97/6.17), m3/M3 (pKi = 6.11/6.04) and m5 sites (pKi = 5.84). Functional studies in vitro showed BIBN 99 to be a competitive antagonist and to have an 11- to 25-fold higher affinity for M2 receptors than for putative M1 receptors in the rabbit vas deferens or M3 receptors in guinea-pig trachea. In vivo studies revealed that BIBN 99 is able to cross the blood-brain barrier, and although showing an approximately 3-fold higher affinity for M2 binding sites BIBN 99 appeared to be 7- to 18-fold less potent than AF-DX 116 in inhibiting muscarinic agonist or vagally induced bradycardia in rats and guinea-pigs. The results show that BIBN 99 is the first lipophilic muscarinic M2 receptor antagonist to have remarkable M2 versus M1 selectivity (30-fold). In addition, BIBN 99 possesses central nervous system activity and only minor peripheral cardiac effects.

Pharmacological characterization of the novel nonpeptide angiotensin II receptor antagonist, BIBR 277.

1. The pharmacological profile of BIBR 277, 4'-[(1,4'-dimethyl-2'-propyl[2,6'-bi-1H-benzimidazol]-1'-yl)methyl ]- [1,1'-biphenyl]-2-carboxylic acid, a novel, nonpeptide angiotensin II receptor antagonist has been investigated by use of receptor binding studies, enzymatic assays, functional in vitro assays in rabbit aorta as well as in vivo experiments in pithed, anaesthetized and conscious rats. 2. BIBR 277 potently interacted with rat AT1 receptors (Ki 3.7 nM). Competitive receptor interaction was shown by radioligand saturation experiments performed in the presence of BIBR 277. The failure to inhibit radioligand binding to AT2 sites demonstrates the selectivity of BIBR 277 for AT1 receptors. This is further substantiated by the findings that BIBR 277 neither interacted with other receptor systems investigated nor affected the activity of components of the human renin-angiotensin system, such as plasma renin or serum converting enzyme. 3. In rabbit aorta, BIBR 277 had no agonistic properties and was shown to be an insurmountable antagonist of angiotensin II-induced contractions (KB 0.33 nM). The antagonistic effect persisted even after several wash-out procedures. However, this interaction was not irreversible since the insurmountable antagonism was concentration-dependently reversed when BIBR 277 (0.1 microM) and the surmountable antagonist, losartan (0.1 and 1.0 microM) were incubated simultaneously. The specificity of BIBR 277 for the AT1 receptor was further substantiated in this preparation since micromolar concentrations of BIBR 277 neither affected potassium chloride and noradrenaline-induced contractions nor acetylcholine-mediated tissue relaxation. 4. In pithed rats, i.v. administration of BIBR 277 (0.1, 0.3 and 1.0 mg kg-1) shifted the dose-pressor response curve to angiotensin II dose-dependently to the right with ED50 values of 0.23 microg kg-1 (control)and 1.4 microg kg-1, 4.7 microg kg-1 and 20 microg kg-1, respectively. As observed in the in vitro experiments no agonistic effect was detected and the maximum of the blood pressure response to angiotensin II at the highest dose of BIBR 277 was decreased by 29%.5. In anaesthetized rats, bolus i.v. administration of 0.1, 0.3 and 1.0 mg kg-1 BIBR 277 attenuated the blood pressure response to bolus i.v. injections of angiotensin 11 (0.1 microg kg-1). At the highest dose an almost complete blockade was observed even after 2 h.6. Single oral administration of BIBR 277 (0.3 and 1.0 mg kg-1) to conscious, chronically instrumented renovascular hypertensive rats dose-dependently decreased the mean arterial blood pressure by 15 and 30 mmHg, respectively. At the higher dose a significant antihypertensive effect was maintained for more than 24 h. Moreover, consecutive daily dosing of 1 mg kg-1 orally resulted in a sustained reduction in blood pressure over the 4 day observation period.7. It is concluded that BIBR 277 is an effective and selective angiotensin II antagonist with antihypertensive activity after oral administration.

Angiotensin II receptor antagonists.

The development of novel non-peptide compounds with high affinity for-angiotensin II (Ang II) receptors has greatly facilitated the subclassification of Ang II receptors into AT1- and AT2-receptor subtypes. Whereas PD 123177 (1-(4-amino-3-methylphenyl)methyl-5-diphenyl-acetyl-4,5,6,7-tetrahydro- 1H-imidazol [4,5-c]pyridine-6-carboxylic acid) is the prototypical antagonist for AT2-receptors, DuP 753 (2-n-butyl-4-chloro-5-hydroxymethyl-1-[(2'-(1 H-tetrazol-5-yl)biphenyl-4-yl) methyl) imidazole, losartan) is the prototypical antagonist for AT1-receptors. So far, non-selective non-peptide Ang II receptor antagonists have not been identified although AT1/AT2 selectivity ratios of 17 and 37 have already been reported for BIBS 39 (4'-[(2-n-butyl-6- cyclohexylaminocarbonylamino-benzimidazole-1-yl)-methyl] biphenyl-2-carboxylic acid) and BIBS 222 (2-n-butyl-1-[4-(6-carboxy-2, 5-dichlorobenzoylamino)-benzyl]-6-N-(methylaminocarbonyl)- n-pentylaminobenzimidazole). Functional studies with AT1-antagonists indicate that Ang II antagonism at the receptor level can be rather complex. Experimental data suggests that not only are receptor binding kinetics involved, but also that additional binding sites, and possibly even AT1 subtypes, are involved. The antihypertensive activity of the AT1-antagonist DuP 753 is demonstrated in a high renin (2K 1C) and a low renin (TGRmREN2) hypertensive rat model. The kidney especially is very sensitive to Ang II and this organ seems to be a target for Ang II receptor antagonists. This can be demonstrated with experiments on the isolated rat kidney.

Therapeutic potential of CNS-active M2 antagonists: novel structures and pharmacology.

Clinical trials with muscarinic agonists or acetylcholine esterase inhibitors for the treatment of Alzheimer's dementia have shown disappointing or equivocal results. An alternative treatment of this disease is the development of drugs which enhance the release of acetylcholine. It is believed, that of the five muscarinic receptor subtypes so far identified in the brain, M2 receptors are located presynaptically in the cortex and hippocampus and upon stimulation inhibit the release of acetylcholine. Based on this hypothesis, we initiated a drug discovery program with the aim of identifying selective and centrally active M2 antagonists which are capable of enhancing cholinergic transmission. These efforts resulted in the successful design and synthesis of novel muscarinic antagonists able to cross the blood brain barrier. Moreover, these compounds show few peripheral effects and possess a superior M2 versus M1 selectivity. The prototype of this novel class of M2 selective compounds, BIBN 99, could be a valuable tool to test the hypothesis that lipophilic M2 antagonists show beneficial effects in the treatment of cognitive disorders.