Laramie Mary Gaster

SB-216641 AND BRL-15572 - COMPOUNDS TO PHARMACOLOGICALLY DISCRIMINATE H5-HT1B AND H5-HT1D RECEPTORS

Abstract Despite only modest homology between h5-HT1B and h5-HT1D receptor amino acid sequences, these receptors display a remarkably similar pharmacology. To date there are few compounds which discriminate between these receptor subtypes and those with some degree of selectivity, such as ketanserin, have greater affinity for other 5-HT receptor subtypes. We now report on two compounds, SB-216641 (N-[3-(2-dimethylamino) ethoxy-4-methoxyphenyl]-2’-methyl-4’-(5-methyl-1,2,4-oxadiazol-3-yl)-(1,1’-biphenyl)-4-carboxamide) and BRL-15572 3-[4-(3-chlorophenyl) piperazin-1-yl]-1,1-diphenyl-2-propanol), which display high affinity and selectivity for h5-HT1B and h5-HT1D receptors, respectively. In receptor binding studies on human receptors expressed in CHO cells, SB-216641 has high affinity (pKi=9.0) for h5-HT1B receptors and has 25-fold lower affinity at h5-HT1D receptors. In contrast, BRL-15572 has 60-fold higher affinity for h5-HT1D (pKi=7.9) than 5-HT1B receptors. Similar affinities for these compounds were determined on native tissue 5-HT1B receptors in guinea-pig striatum. Functional activities of SB-216641 and BRL-15572 were measured in a [35S]GTPγS binding assay and in a cAMP accumulation assay on recombinant h5-HT1B and h5-HT1D receptors. Both compounds were partial agonists in these high receptor expression systems, with potencies and selectivities which correlated with their receptor binding affinities. In the cAMP accumulation assay, results from pKB measurements on the compounds again correlated with receptor binding affinities (SB-216641, pKB=9.3 and 7.3; BRL-15572, pKB=<6 and 7.1, for h5-HT1B and h5-HT1D receptors respectively). These compounds will be useful pharmacological agents to characterise 5-HT1B and 5-HT1D receptor mediated responses.

Effects of selective h5-HT1B (SB-216641) and h5-HT1D (BRL-15572) receptor ligands on guinea-pig and human 5-HT auto- and heteroreceptors.

Human cerebral cortical slices and synaptosomes, guinea-pig cerebral cortical slices and human right atrial appendages were used to study the effects of SB-216641, a preferential h5-HT1B receptor ligand, and of BRL-15572, a preferential h5-HT1D receptor ligand, on the presynaptic h5-HT1B and h5-HT1B-like autoreceptors in the human and guinea-pig brain preparations, respectively, and on the presynaptic h5-HT1D heteroreceptors in the human atrium. The brain preparations, preincubated with [3H]serotonin ([3H]5-HT), and the segments of atrial appendages, preincubated with [3H]noradrenaline, were superfused with modified Krebs’ solution and tritium overflow was evoked electrically (human and guinea-pig cerebral cortex slices and human atrial appendages) or by high K+ (human cerebral cortex synaptosomes). The electrically evoked tritium overflow from guinea-pig cerebral cortex slices was reduced by the 5-HT receptor agonist 5-carboxamidotryptamine (5-CT). This effect was not modified by BRL-15572 (2μM; concentration 154 times higher than its Ki at h5-HT1D receptors) but was antagonized by SB-216641 (0.1μM; concentration 100 times higher than its Ki at h5-HT1B receptors; apparent pA2 8.45). SB-216641 (0.1μM) by itself facilitated, whereas BRL-15572 (2μM) did not affect, the evoked overflow. In human cerebral cortex slices SB-216641 (0.1μM) also facilitated, and BRL-15572 (2μM) again failed to affect, the electrically evoked tritium overflow. In human cerebral cortical synaptosomes, 5-CT reduced the K+-evoked tritium overflow. This response was unaffected by BRL-15572 (300nM) but antagonized by SB-216641 (15nM; drug concentrations 23 and 15 times higher than their Ki at h5-HT1D and h5-HT1B receptors, respectively). Both drugs, given alone, did not modify the K+-evoked tritium overflow. In human atrial appendages, the electrically evoked tritium overflow was inhibited by 5-HT in a manner susceptible to antagonism by BRL-15572 (300nM; 23 times Ki at h5-HT1D receptors) but not by SB-216641 (30nM; 30 times Ki at h5-HT1B receptors). Both drugs by themselves did not change the electrically evoked tritium overflow. In conclusion, SB-216641 behaves as a preferential antagonist at native human 5-HT1B receptors and BRL-15572 as a preferential antagonist at native human 5-HT1D receptors. These compounds are clearly useful tools for the differentiation between human 5-HT1B and 5-HT1D receptors in functional studies.

Increased defaecation caused by 5-HT4 receptor activation in the mouse

The precursor to 5-hydroxytryptamine (5-HT), 5-hydroxytryptophan, (5-HTP, 5-50 mg.kg-1) administered subcutaneously (s.c.) to conscious, fed mice caused a dose dependent increase in faecal pellet and fluid output. To avoid provoking watery diarrhoea, all experiments were performed using 5-HTP at 10 mg.kg-1. This dose caused maximal increases in the fluid content (471 +/- 41%) and number of formed faecal pellets defaecated (328 +/- 13% n = 25), 10 and 20 min respectively after administration, when compared to saline-treated mice. In both saline- and 5-HTP-treated mice methiothepin, ketanserin, mianserin and granisetron reduced defaecation at high s.c. doses (100 micrograms.kg-1 or 1000 micrograms.kg-1). The 5-HT4 receptor antagonists, DAU 6285 (endo-6-methoxy-8-methyl-8-azabicyclo[3.2.1]oct-3-yl-2,3-dihydro-2-oxo-1 H-benzimidazole-1-carboxylate hydrochloride), SDZ 205-557 (2-methoxy-4-amino-5-chloro-benzoic acid 2-(diethylamino) ethyl ester) and SB 204070 ([1-butyl-4-piperidinylmethyl]-8-amino-7-chloro-1,4-benzodioxan -5- carboxylate), had no effects when administered s.c. to saline-treated mice, but dose-dependently inhibited the 5-HTP-evoked responses. Only SB 204070 at 1000 micrograms.kg-1 completely inhibited the responses to 5-HTP returning them to normal levels. We conclude that SB 204070 is a potent antagonist for the investigation of 5-HT4 receptor function in both normal and disturbed gastrointestinal activity.

Blockade of human atrial 5-HT4 receptors by SB 207710, a selective and high affinity 5-HT4 receptor antagonist

The mode of antagonism of 5-hydroxytryptamine-induced positive inotropic effects by the highly selective 5-HT4 receptor antagonist SB 207710 (1-butyl4-piperidinyl) methyl 8-amino-7-iodo-1,4-benzodioxan-5-carboxylate was investigated on isolated preparations of human right atrial appendage. SB 207 710 caused concentration-dependent (0.1–10 nmol/l) surmountable antagonism of the effects of 5-hydroxytryptamine with a pKB (mol/l) of 10.1. Due to its high selectivity and affinity, SB 207710 could be a powerful tool for the comparison of human atrial 5-HT4 receptors with 5-HT4 receptors of other organs of man and other species.

Quaternised renzapride as a potent and selective 5-HT4 receptor agonist

The n-butyl quaternary salt of the 5-HT4 receptor agonist and 5-HT3 receptor antagonist, renzapride, is reported to be a potent and selected 5-HT4 receptor agonist.

Azabicyclic indole esters as potent 5-HT4 receptor antagonists.

The synthesis of a series of azabicyclic indole esters is described and their potency reported as 5-HT4 receptor antagonists. Optimization of the most potent compound (19) by preparing the corresponding oxazino[3,2-a]indole ester afforded 34, which had a pIC50 of 9.5 in the guinea pig distal colon longitudinal muscle myenteric plexus preparation.

5-HT4receptor antagonists : oxazolo, oxazino and oxazepino[3,2-a]indole derivatives

Abstract The identification of oxazolo, oxazino and oxazepino[3,2-a]indoles as new structural classes of highly potent 5-HT4 receptor antagonists is described. Compounds (4) (5) and (6) are among the most potent 5-HT4 receptor antagonists reported to date.

Synthesis and 5-HT3 receptor antagonist potency of novel (endo) 3,9-diazabicyclo[3.3.1]nonan-7-amino derivatives

Abstract The synthesis and 5-HT 3 receptor affinity and/or antagonist activity of a series of novel N-(3,9-diazabicyclo[3.3.1]nonan-7-yl)amides (3-azagranatanes) are described. High potency is retained with relatively bulky 3-N substituents indicative of a large sterically allowed volume “underneath” the piperidine ring.