In vitro alpha1-adrenoceptor pharmacology of Ro 70-0004 and RS-100329, novel alpha1A-adrenoceptor selective antagonists.

It has been hypothesized that in patients with benign prostatic hyperplasia, selective antagonism of the alpha1A-adrenoceptor-mediated contraction of lower urinary tract tissues may, via a selective relief of outlet obstruction, lead to an improvement in symptoms. The present study describes the alpha1-adrenoceptor (alpha1-AR) subtype selectivities of two novel alpha1-AR antagonists, Ro 70-0004 (aka RS-100975) and a structurally-related compound RS-100329, and compares them with those of prazosin and tamsulosin. Radioligand binding and second-messenger studies in intact CHO-K1 cells expressing human cloned alpha1A-, alpha1B- and alpha1D-AR showed nanomolar affinity and significant alpha1A-AR subtype selectivity for both Ro 70-0004 (pKi 8.9: 60 and 50 fold selectivity) and RS-100329 (pKi 9.6: 126 and 50 fold selectivity) over the alpha1B- and alpha1D-AR subtypes respectively. In contrast, prazosin and tamsulosin showed little subtype selectivity. Noradrenaline-induced contractions of human lower urinary tract (LUT) tissues or rabbit bladder neck were competitively antagonized by Ro 70-0004 (pA2 8.8 and 8.9), RS-100329 (pA2 9.2 and 9.2), tamsulosin (pA2 10.4 and 9.8) and prazosin (pA2 8.7 and 8.3 respectively). Affinity estimates for tamsulosin and prazosin in antagonizing alpha1-AR-mediated contractions of human renal artery (HRA) and rat aorta (RA) were similar to those observed in LUT tissues, whereas Ro 70-0004 and RS-100329 were approximately 100 fold less potent (pA2 values of 6.8/6.8 and 7.3/7.9 in HRA/RA respectively). The alpha1A-AR subtype selectivity of Ro 70-0004 and RS-100329, demonstrated in both cloned and native systems, should allow for an evaluation of the clinical utility of a 'uroselective' agent for the treatment of symptoms associated with benign prostatic hyperplasia.     

Design and synthesis of novel alpha(1)(a) adrenoceptor-selective antagonists. 1. Structure-activity relationship in dihydropyrimidinones

Dihydropyrimidinones such as compound 12 exhibited high binding affinity and subtype selectivity for the cloned human alpha(1a) receptor. Systematic modifications of 12 led to identification of highly potent and subtype-selective compounds such as (+)-30 and (+)-103, with high binding affinity (K(i) = 0.2 nM) for alpha(1a) receptor and greater than 1500-fold selectivity over alpha(1b) and alpha(1d) adrenoceptors. The compounds were found to be functional antagonists in human, rat, and dog prostate tissues. Compound (+)-103 exhibited excellent selectively to inhibit intraurethral pressure (IUP) as compared to lowering diastolic blood pressure (DBP) in mongrel dogs (K(b)(DBP)/K(b)(IUP) = 40) suggesting uroselectivity for alpha(1a)-selective compounds.     

Alpha1-adrenoceptor antagonists. 6. Structural optimization of pyridazinone-arylpiperazines. Study of the influence on affinity and selectivity of cyclic substituents at the pyridazinone ring and alkoxy groups at the arylpiperazine moiety

In continuing our search for selective alpha(1)-adrenoceptor (AR) antagonists, we have synthesized new alkoxyarylpiperazinylalkylpyridazinone derivatives. The new compounds were tested for their affinity toward alpha(1)- and alpha(2)-AR and toward the 5-HT(1A) receptor. alpha(1)-AR affinity data are in the subnanomolar range, with 3 showing an affinity of 0.052 nM, about 5-fold higher than prazosin. None of the studied compounds was found to be alpha(1)/alpha(2) selective, but 8 showed an interesting 5-HT(1A)/alpha(1) affinity ratio of 119.     

Evaluation of the pharmacological selectivity profile of alpha 1 adrenoceptor antagonists at prostatic alpha 1 adrenoceptors: binding, functional and in vivo studies.

1. The profile of a range of alpha 1 adrenoceptor antagonists was determined in vitro against cloned human alpha 1A, alpha 1B and alpha 1D adrenoceptors and against noradrenaline-mediated contractions of rat aorta and human prostate. The in vivo profile of compounds was determined in an anaesthetized dog model which allowed the simultaneous assessment of antagonist potency against phenylephrine-mediated increases in blood pressure and prostatic pressure. 2. The quinazoline antagonists, prazosin, doxazosin and alfuzosin displayed high affinity but were non selective for the three cloned human alpha 1 adrenoceptors. Indoramin and SNAP 1069 showed selectivity for alpha 1A and alpha 1B adrenoceptors relative to the alpha 1D subtype. Rec 15/2739, WB 4101, SL 89,0591, (+)- and (-)- tamsulosin showed selectivity for alpha 1A and alpha 1D adrenoceptors relative to the alpha 1B subtype. RS 17053 showed high affinity and selectivity for alpha 1A adrenoceptors (pKi 8.6) relative to alpha 1B (pKi = 7.3) and alpha 1D (pKi = 7.1) subtypes. 3. (+)-Tamsulosin, (-)-tamsulosin, SL 89,0591, Rec 15/2739, SNAP 1069 and RS 17053 appeared to act as competitive antagonists of noradrenaline-mediated contractions of rat aorta yielding pA2 affinity estimates which were similar to binding affinities at cloned human alpha 1D adrenoceptors. The following rank order was obtained: prazosin = (-)-tamsulosin > doxazosin > SL 89,0591 = (+)-tamsulosin > Rec 15/2739 > RS 17053 = SNAP 1069. 4. (-)-Tamsulosin was a very potent, insurmountable antagonist of noradrenaline-mediated contractions of human prostate, yielding an approximate pA2 estimate of 9.8 at 1 nM. The corresponding (+)-enantiomer was 30 fold weaker. SL 89,0591, SNAP 1069 and Rec 15/2739 yielded pA2 estimates which compared well with their alpha 1A binding affinities. The affinity estimate for prazosin on human prostate was lower than the corresponding binding affinity determined at alpha 1A adrenoceptors and RS 17053 was a very weak antagonist on human prostate (pA2 = 6.0) relative to the high affinity (pKi = 8.6) determined at cloned human alpha 1A adrenoceptors. 5. In the anaesthetized dog, in vivo pseudo "pA2' values showed that doxazosin, (+)- and (-)-tamsulosin inhibited phenylephrine-induced increases in prostatic and blood pressure with similar affinity, implying that these agents show little or no selectivity for prostatic responses in this model. SL 89,0591 and SNAP 1069 were moderately selective (3 and 6 fold respectively) for prostatic pressure relative to blood pressure. Rec 15/2739 was a more potent antagonist of phenylephrine-mediated increases in prostatic pressure ("pA2' = 8.74) compared to blood pressure ("pA2' = 7.51). 6. Data in this study suggest that the alpha 1 adrenoceptor mediating noradrenaline-induced contractions of human prostate, whilst having some of the characteristics of an alpha 1A adrenoceptor, cannot be satisfactorily aligned with cloned alpha 1A, alpha 1B or alpha 1D adrenoceptors. In addition, studies in the anaesthetized dog have shown that agents having high affinity and selectivity for prostatic alpha 1 adrenoceptors, particularly over the alpha 1D subtype, appear to inhibit phenylephrine-induced increases in prostatic pressure selectively compared to blood pressure.     

Alpha(1A)-adrenoceptors mediate contractions to phenylephrine in rabbit penile arteries

BACKGROUND AND PURPOSE: Maintained penile erection depends on the absence of alpha-adrenoceptor (alpha-AR) activation and so can be facilitated by alpha-blockers. This study seeks the alpha(1)-AR subtypes involved in order to inform the pro-erectile consequences of subtype selective blockade. EXPERIMENTAL APPROACH: Wire myography was used with dorsal (nutritional supply) and cavernous (erectile inflow) penile arteries; standard alpha-AR-selective agonists and antagonists were employed to classify responses. KEY RESULTS: In both penile arteries noradrenaline (NA) and phenylephrine (PE, alpha(1)-AR agonist) caused concentration-dependent contractions. Sensitivity to NA was increased by NA uptake blockers, cocaine (3 microM) and corticosterone (30 microM). PE responses were antagonised by phentolamine (non-selective alpha-AR: dorsal pK(B) 8.00, cavernous 8.33), prazosin (non-subtype-selective alpha(1)-AR: dorsal 8.60, cavernous 8.41) and RS100329 (alpha(1A)-AR selective: dorsal 9.03, cavernous 8.80) but not by BMY7378 (alpha(1D)-AR selective: no effect at 1-100 nM) or Rec15/2615 (alpha(1B)-AR selective: no effect at 1-100 nM). Schild analysis was straightforward in cavernous artery, indicating that PE activates only alpha(1A)-AR. In dorsal artery Schild slopes were low, though alpha(1A)-AR was still indicated. Analysis using UK 14,304 and rauwolscine indicated an alpha(2)-AR component in dorsal artery that may account for low slopes to alpha(1)-AR antagonists. CONCLUSIONS AND IMPLICATIONS: Penile arteries have a predominant, functional alpha(1A)-AR population with little evidence of other alpha(1)-AR subtypes. Dorsal arteries (nutritional supply) also have alpha(2)-ARs. Thus, alpha-AR blockers with affinity for alpha(1A)-AR or alpha(2)-AR would potentially have pro-erectile properties; the combination of these perhaps being most effective. This should inform the design of drugs to assist/avoid penile erection.     

Characterization of an alpha 1D-adrenoceptor mediating the contractile response of rat aorta to noradrenaline.

1. The affinities of a number of alpha 1-adrenoceptor antagonists were determined by displacement of [3H]-prazosin binding from cloned human alpha 1A-adrenoceptors (previously designated cloned alpha 1c subtype), alpha 1B alpha 1D and rat alpha 1D-adrenoceptors, stably expressed in rat-1 fibroblasts. Functional affinity estimates for these compounds were also determined from noradrenaline-mediated contractions of rat aorta. 2. BMY 7378 displayed high affinity for cloned human alpha 1D-adrenoceptors (pKi = 8.2 +/- 0.10) and was selective over alpha 1A (pKi = 6.2 +/- 0.10) and alpha 1B subtypes (6.7 +/- 0.11). WB 4101, benoxathian and phentolamine displayed high affinity for alpha 1A and alpha 1D adrenoceptors compared to the alpha 1B subtype. Spiperone displayed high affinity and selectivity for alpha 1B adrenoceptors (pKi 8.8 +/- 0.16). 5-Methyl-urapidil was selective for cloned alpha 1A adrenoceptors. 3. Comparative binding affinities (pKi) for compounds at cloned human and rat1D adrenoceptors were almost identical (r = 0.99, slope = 1.08). 4. Prazosin, doxazosin and 5-methyl-urapidil were potent, competitive antagonists of noradrenaline-mediated contractions of rat aorta (pA2 values of 9.8, 8.8 and 7.8 respectively). The selective alpha 1D antagonist BMY 7378 was also a potent antagonist on rat aorta (pKB = 8.3 +/- 0.1) but the interaction of this compound was not consistent with competitive antagonism at a single population of receptors. 5. Functional affinities for compounds determined against noradrenaline-mediated contractions of rat aorta correlated well with binding affinities at cloned alpha 1D-adrenoceptors (r = 0.96), but not with alpha 1A (r = 0.61) or alpha 1B (r = 0.46) subtypes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pharmacological properties of naftopidil, a drug for treatment of the bladder outlet obstruction for patients with benign prostatic hyperplasia

Naftopidil, a phenylpiperazine derivative, is a novel alpha 1-adrenoceptor antagonist and is new drug for the bladder outlet obstruction in patients with benign prostatic hyperplasia (BPH). Naftopidil competitively inhibited specific [3H]prazosin binding in prostatic membranes of humans, and its Ki value was 11.6 nM. Using cloned human alpha 1-adrenoceptor subtypes (alpha 1a, alpha 1b and alpha 1d), naftopidil was selective for the alpha 1d-adrenoceptor with approximately 3- and 17-fold higher affinity than for the alpha 1a- and alpha 1b-adrenoceptor subtypes, respectively. In anesthetized dogs, naftopidil selectively inhibited the phenylephrine-induced increase in prostatic pressure compared with mean blood pressure. The selectivity of naftopidil for prostatic pressure was more potent than those of tamsulosin and prazosin. In conscious rabbits, the effect of naftopidil on the blood pressure reactions following the tilting was less potent than those of tamsulosin and prazosin. In clinical studies, naftopidil has been demonstrated to be effective in the treatment of bladder outlet obstruction in patients with BPH. In Japan, naftopidil has been already approved for clinical use as a drug for BPH.     

Bradykinin agonist activity of a novel, potent oxytocin antagonist

From a series of potent cyclic hexapeptide oxytocin (OT) antagonists, a compound that exhibited significant bradykinin (BK) agonist activity was identified. L-366,811 (cyclo[L-proline-D-tryptophan-L-isoleucine-D-pipecolic acid-L-piperazine-2-carboxylic acid-N-Me-D-phenylalanine]) stimulated phosphatidylinositol (PI) turnover in rat uterine slices in vitro (approximately EC50, 2 microM) with a maximal effect (15-fold increase over basal) greater than that obtained for either BK or OT. L-366,811 also elicited dose-related contractions of the isolated rat uterus, producing measurable effects at 100 nM. Several other equally potent OT antagonists from the cyclic hexapeptide structural class were either less potent or inactive as activators of uterine PI turnover or contractility. The stimulatory effects of L-366,811 on uterine PI turnover and contractions were blocked by BK antagonists but not by an arginine vasopressin (AVP)/OT antagonist. In radioligand binding studies, L-366,811 exhibited moderate affinity (IC50, 360 nM) for the [3H]BK binding site in rat uterus, consistent with its potency in the functional models. These results indicate that L-366,811 exhibits BK agonist activity in rat uterus in vitro.     

Amino acids of the human alpha1d-adrenergic receptor involved in antagonist binding

Computer simulations of the human alpha(1d)-adrenergic receptor (alpha(1d)-AR) based on the crystal structure of rhodopsin have been combined with experimental site-directed mutagenesis to investigate the role of residues in the transmembrane domains in antagonist binding. Our results indicate that the amino acids Asp176 in the third transmembrane domain (TMD), Glu237 in TMD IV, and Ser258 in TMD V of alpha(1d)-AR were directly involved in prazosin and tamsulosin binding. The Asp176Ala mutant did not exhibit any affinity for [(3)H]prazosin and neither did it show agonist-stimulated inositol phosphates (IP) formation. On the other hand, the Glu237Ala and Ser258Ala mutant alpha(1d)-AR showed increased binding affinity for [(3)H]prazosin. Competition binding experiments showed that prazosin affinity had increased to 5-fold and 3-fold in the Glu237Ala and Ser258Ala mutants, respectively, versus wild-type; and tamsulosin affinity only increased in the Ser258Ala mutant (2-fold vs wild-type). It seems that these two residues constrain the receptor by interaction with other residues and this disruption of the interaction increased the receptor's binding affinity towards antagonists. However, the Glu237Ala and Ser258Ala mutant receptors retained the ability to stimulate the formation of myo-[(3)H]inositol but had activities lower than that of the wild-type receptor. The present results provide direct evidence that these amino acid residues are responsible for the interactions between alpha(1d)-AR and the radioligand [(3)H]prazosin as well as tamsulosin.     

Effect of tamsulosin on spontaneous bladder contraction in conscious rats with bladder outlet obstruction: comparison with effect on intraurethral pressure

We investigated the effect of tamsulosin, an alpha(1)-adrenoceptor antagonist, on bladder function, especially spontaneous bladder contractions before micturition (premicturition contraction), in conscious rats with bladder outlet obstruction induced by partial urethral ligation, and compared the results with the effect on intraurethral pressure response in anesthetized rats. In obstructed rats, the alpha(1)-adrenoceptor antagonists tamsulosin, naftopidil and urapidil and non-selective alpha-adrenoceptor antagonist phentolamine inhibited premicturition contractions in a dose-dependent fashion. In contrast, yohimbine, an alpha(2)-adrenoceptor antagonist, and atropine, a muscarinic receptor antagonist, hardly inhibited them. Tamsulosin and urapidil showed clearly inhibitory effects on increases in intraurethral pressure induced by phenylephrine, an alpha(1)-adrenoceptor agonist, in the same dose range as that at which they inhibited premicturition contractions, whereas naftopidil required somewhat higher doses to inhibit increases in intraurethral pressure than those at which it inhibited premicturition contractions. In conclusion, premicturition contractions observed in obstructed rats were sensitive to alpha(1)-adrenoceptor antagonists, but not to alpha(2)-adrenoceptor or muscarinic receptor antagonists. Tamsulosin was shown to be effective against both premicturition contraction and intraurethral pressure response in the same dose range in rats. These results partly support the fact that tamsulosin has improved storage symptoms as well as voiding symptoms in patients with lower urinary tract symptoms associated with bladder outlet obstruction by blocking alpha(1)-adrenoceptors.     

In vitro and in vivo evaluation of dihydropyrimidinone C-5 amides as potent and selective alpha(1A) receptor antagonists for the treatment of benign prostatic hyperplasia

alpha(1) Adrenergic receptors mediate both vascular and lower urinary tract tone, and alpha(1) receptor antagonists such as terazosin (1b) are used to treat both hypertension and benign prostatic hyperplasia (BPH). Recently, three different subtypes of this receptor have been identified, with the alpha(1A) receptor being most prevalent in lower urinary tract tissue. This paper explores 4-aryldihydropyrimidinones attached to an aminopropyl-4-arylpiperidine via a C-5 amide as selective alpha(1A) receptor subtype antagonists. In receptor binding assays, these types of compounds generally display K(i) values for the alpha(1a) receptor subtype <1 nM while being greater than 100-fold selective versus the alpha(1b) and alpha(1d) receptor subtypes. Many of these compounds were also evaluated in vivo and found to be more potent than terazosin in both a rat model of prostate tone and a dog model of intra-urethral pressure without significantly affecting blood pressure. While many of the compounds tested displayed poor pharmacokinetics, compound 48 was found to have adequate bioavailability (>20%) and half-life (>6 h) in both rats and dogs. Due to its selectivity for the alpha(1a) over the alpha(1b) and alpha(1d) receptors as well as its favorable pharmacokinetic profile, 48 has the potential to relieve the symptoms of BPH without eliciting effects on the cardiovascular system.     

Asp125 and Thr130 in transmembrane domain 3 are major sites of alpha1b-adrenergic receptor antagonist binding

Site-directed mutagenesis was used to investigate the molecular interactions involved in prazosin binding to the human alpha(1b)-adrenergic receptor (alpha(1b)-AR) receptor. Based on molecular modeling studies, Thr130 and Asp125 in transmembrane region III of the alpha(1b)-AR receptor were found to interact with prazosin. Thr130 and Asp125 were mutated to alanine (Ala) and expressed in HEK293 cells. The radioligand [(3)H]prazosin did not show any binding to Asp125Ala mutant of alpha(1b)-AR. Therefore, it was not possible to find any prazosin affinity to the mutant using the radioligand [(3)H]prazosin. The mutation also abolished phenylephrine-stimulated inositol phosphate (IP) formation of [(3)H]myo-inositol. On the other hand, the Thr130Ala mutant showed reduced binding affinity for [(3)H]prazosin (dissociation constant, K(d) 674.27 pM versus 90.27 pM for the wild-type receptor) and had reduced affinity for both tamsulosin and prazosin (11-fold and 9-fold, respectively). However, the Thr130Ala mutant receptor retained the ability to stimulate the formation of [(3)H]myo-inositol. The results provide direct evidence that Asp125 and Thr130 are responsible for the interactions between alpha(1b)-AR receptor and radioligand [(3)H]prazosin as well as tamsulosin.     

Synthesis and structure-activity relationships of 2-sulfonamido-1,3,4,6,7,11b alpha-hexahydro-2H-benzo[a]quinolizines as alpha 2-adrenoceptor antagonists

A series of 2-sulfonamido-1,3,4,6,7,11b alpha-hexahydro-2H-benzo[a]quinolizines were synthesized and examined for alpha 2- and alpha 1-adrenoceptor antagonist activity on the rat vas deferens and anococcygeus muscle, respectively. A number of compounds in this series were shown to be potent and selective alpha 2-adrenoceptor antagonists. Studies on the resolved enantiomers of compounds 6, 10, and 16 showed that alpha 2-adrenoceptor antagonist activity resided primarily in the 2R,11bS isomers, related to the absolute configuration of the alpha 2-antagonist yohimbine, such that the benzene ring and sulfonamide groups in this series were superimposable on the pyrrole and ester groups of yohimbine.     

Vascular activity of (-)-anonaine, (-)-roemerine and (-)-pukateine, three natural 6a(R)-1,2-methylenedioxyaporphines with different affinities for alpha1-adrenoceptor subtypes

We have studied the mechanism of action of three 6a( R)-1,2-methylenedioxyaporphines as vasorelaxant compounds. The alkaloids assayed showed different affinities for the three human cloned alpha (1)-adrenoceptor (AR) subtypes stably expressed in rat-1 fibroblasts, showing lower affinity for alpha(1B)-AR with regard to the alpha(1A)- or alpha(1D)-subtypes. These three natural compounds are more potent inhibitors of [ (3)H]-prazosin binding than of [ (3)H]-diltiazem binding to rat cerebral cortical membranes. As all these alkaloids inhibited noradrenaline (NA)-induced [ (3)H]-inositol phosphate formation in cerebral cortex and rat tail artery, they may be safely viewed as alpha (1)-AR antagonists, as is demonstrated by the vasorelaxant responses observed in isolated rat tail artery and/or aorta precontracted with NA. The alkaloids also inhibited the contractile response evoked by KCl (80 mM) but with a lower potency than that shown against NA-induced contraction. We have also examined their ability to inhibit the different forms of cyclic nucleotide phosphodiesterases (PDE) isolated from bovine aortic smooth muscle and endothelial cells, with negative results. We conclude that N-methylation favours the interaction of (R)-aporphines with all alpha (1)-AR subtypes, and that the topography of the binding site recognizing the basic or protonated nitrogen atom is similar in all three alpha (1)-AR subtypes. The presence of a hydroxy group at C-11 has different effects on the affinity for each alpha (1)-AR subtype but decreases the affinity for Ca (2+) channels. These results confirm and extend the view that subtle changes in the hydroxylation patterns on the aromatic ring of the aporphine structure affect the interactions of these compounds with the three alpha (1)-AR subtypes in different ways, suggesting that the binding site recognizing the aporphine skeleton is different in each of the three subtypes.     

Enhancement of apomorphine-induced penile erection in the rat by a selective alpha(1D)-adrenoceptor antagonist

1. Effects of A-322312 (alpha(1B)-adrenoceptor (AR) antagonist), A-119637 (alpha(1D)-AR antagonist), prazosin (non-selective alpha(1)-AR antagonist), and yohimbine (alpha(2)-AR antagonist) were studied in rat corpus cavernosum (CC) and cavernous artery (Acc) preparations. Effects of intracavernous (i.c.) or intraperitoneal (i.p.) administration of alpha(1)-AR antagonists on apomorphine-induced erections were investigated. 2. A-119637 attenuated electrically induced contractions in isolated CC (-logIC(50); 8.12+/-0.15), and relaxed noradrenaline (NA)-contracted preparations by more than 90% at 10(-7) M. At the same concentration, the -logEC(50) value for NA in Acc was altered from 6.79+/-0.07 to 4.86+/-0.13. In the CC and Acc, prazosin similarly inhibited contractile responses. 3. Inhibitory effects of A-322312 (10(-7) M) in electrically activated CC were 32.3+/-5.1%, whereas no effect on concentration-response curves for NA was observed in the Acc. Yohimbine (10(-8) M and 10(-7) M), enhanced electrically-induced contractions in isolated CC by 20 to 50%. At 10(-6) M, inhibitory effects of yohimbine were obtained. 4. A-119637 (0.3 micromol kg(-1), i.p.) tripled the number of erections, and produced a 6 fold increase in the duration of apomorphine-induced erectile responses. A-322312, prazosin, or yohimbine did not enhance erections induced by apomorphine. None of the alpha(1)-AR antagonists significantly increased ICP upon i.c. administration. Decreases in blood pressure were seen with A-119637 and prazosin. 5. The present findings show that there is a functional predominance of the alpha(1D)-AR subtype in the rat erectile tissue, and that blockade of this receptor facilitates rat penile erection induced by a suboptimal dose of apomorphine.     

Design, synthesis, and structure-activity relationships of acetylene-based histamine H3 receptor antagonists

New, potent, and selective histamine H3 receptor antagonists have been synthesized by employing the use of (1) an appropriately positioned nonpolar acetylene spacer group, (2) either a two-carbon straight chain linker or a conformationally restricting trans-cyclopropane ring between the C-4 position of an imidazole headgroup and the acetylene spacer, and (3) a Topliss operational scheme for side-chain substitution for optimizing the hydrophobic domain. Compounds 9-18 are examples synthesized with the two-carbon straight chain linker, whereas 26-31 are analogues prepared by incorporation of the trans-(+/-)-cyclopropane at the C-4 position of an imidazole headgroup. Synthesis of both the (1R,2R)- and (1S, 2S)-cyclopropyl enantiomers of the most potent racemic compound 31 (Ki = 0.33 +/- 0.13 nM) demonstrated a stereopreference in H3 receptor binding affinity for the (1R,2R) enantiomer 32 (Ki = 0.18 +/- 0.04 nM) versus the (1S,2S) enantiomer 33 (Ki = 5.3 +/- 0.5 nM). (1R,2R)-4-(2-(5,5-Dimethylhex-1-ynyl)cyclopropyl)imidazole (32) is one of the most potent histamine H3 receptor antagonists reported to date.     

Identification of a novel 4-aminomethylpiperidine class of M3 muscarinic receptor antagonists and structural insight into their M3 selectivity

Identification of a novel class of potent and highly selective M(3) muscarinic antagonists is described. First, the structure-activity relationship in the cationic amine core of our previously reported triphenylpropionamide class of M(3) selective antagonists was explored by a small diamine library constructed in solid phase. This led to the identification of M(3) antagonists with a novel piperidine pharmacophore and significantly improved subtype selectivity from a previously reported class. Successive modification on the terminal triphenylpropionamide part of the newly identified class gave 14a as a potent M(3) selective antagonist that had >100-fold selectivity versus the M(1), M(2), M(4), and M(5) receptors (M(3): K(i) = 0.30 nM, M(1)/M(3) = 570-fold, M(2)/M(3) = 1600-fold, M(4)/M(3) = 140-fold, M(5)/M(3) = 12000-fold). The possible rationale for its extraordinarily higher subtype selectivity than reported M(3) antagonists was hypothesized by sequence alignment of multiple muscarinic receptors and a computational docking of 14a into transmembrane domains of M(3) receptors.     

Tamsulosin: assessment of affinityof (3)H-P razosin binding to two alpha-1- adrenoceptor subtypes in the canine aorta.

This study was performed to assess the affinity of tamsulosin to the alpha(1L)- in addition to alpha(1B)-adrenoceptor (alpha(1)-AR) subtypes coexisting in the canine aorta using the radioligand binding assay. The antagonistic effects of this drug on contraction of the rat aorta were also assessed, and the results were compared with those obtained with prazosin, amosulalol, labetalol, ketanserin, clonidine and propranolol. The pKi value of tamsulosin to the alpha(1L)-subtype was lower than those of prazosin and HV-723, but higher than those of amosulalol, ketanserin and labetalol. The pKi value of tamsulosin for the alpha(1B)-subtype in the canine aorta was similar to that of prazosin. However, this drug showed a higher pKi value than amosulalol, HV-723, labetalol and ketanserin. On the other hand, the order of inhibition potencies for contraction of the rat aorta by phenylephrine was as follows: prazosin > tamsulosin > amosulalol > HV-723 > labetalol > ketanserin > clonidine > propranolol. Thus, although the affinity of tamsulosin to the alpha(1B)-AR subtype in the canine aorta was as high as that in the bovine prostate reported in our previous study, the affinity (pKi 7. 87) of this drug to alpha(1L)-AR in the canine aorta was lower than that (pKi 8.99) in the bovine prostate. These observations suggested that the pharmacological potencies of tamsulosin in the aorta and prostate may be different.     

Discovery and pharmacological evaluation of growth hormone secretagogue receptor antagonists

The discovery and pharmacological evaluation of potent, selective, and orally bioavailable growth hormone secretagogue receptor (GHS-R) antagonists are reported. Previously, 2,4-diaminopyrimidine-based GHS-R antagonists reported from our laboratories have been shown to be dihydrofolate reductase (DHFR) inhibitors. By comparing the X-ray crystal structure of DHFR docked with our GHS-R antagonists and GHS-R modeling, we designed and synthesized a series of potent and DHFR selective GHS-R antagonists with good pharmacokinetic (PK) profiles. An amide derivative 13d (Ca2+ flux IC50 = 188 nM, [brain]/[plasma] = 0.97 @ 8 h in rat) showed a 10% decrease in 24 h food intake in rats, and over 5% body weight reduction after 14-day oral treatment in diet-induced obese (DIO) mice. In comparison, a urea derivative 14c (Ca2+ flux IC50 = 7 nM, [brain]/[plasma] = 0.0 in DIO) failed to show significant effect on food intake in the acute feeding DIO model. These observations demonstrated for the first time that peripheral GHS-R blockage with small molecule GHS-R antagonists might not be sufficient for suppressing appetite and inducing body weight reduction.     

Synthesis, structure-activity relationship, and receptor pharmacology of a new series of quinoline derivatives acting as selective, noncompetitive mGlu1 antagonists

We describe the discovery and the structure-activity relationship of a new series of quinoline derivatives acting as selective and highly potent noncompetitive mGlu1 antagonists. We first identified cis-10 as a fairly potent mGlu1 antagonist (IC(50) = 20 nM) in a cell-based signal transduction assay on the rat mGlu1 receptor expressed in CHO-K1 cells, and then we were able to design and synthesize highly potent compounds on both rat and human mGlu1 receptors as exemplified by compound cis-64a, which has an antagonist potency of 0.5 nM for the human mGlu1 receptor. We briefly present and discuss the in vitro metabolic stability of the compounds in human liver microsomes. We finally report the pharmacokinetic properties of our lead compound cis-64a.