A number of benzoates derived from 4-amino-5-chloro-2-methoxybenzoic acid and substituted 1-piperidineethanol were synthesized and found to be potent 5-HT4 receptor agonists in the electrically-stimulated myenteric plexus and longitudinal muscle of the guinea pig ileum and the rat esophagus muscle. Monosubstitution of the piperidine ring with Me, OH, NH-Ac, or CONH2 groups gave compounds equipotent to 7a (ML 10302), a 5-HT4 receptor agonist previously reported to have nanomolar affinity. 7a,k were as potent as serotonin (5-HT) but had maximal responses which were only 60-80% of that of 5-HT, suggesting a partial agonist profile for these compounds. Binding assays were performed with [3H]GR 113808 in the rat striatum, and several of these compounds were found to have nanomolar affinity for 5-HT4 receptors (7a, Ki = 1.07 +/- 0.5 nM; 7k, Ki = 1.0 +/- 0.3 nM). The introduction of two methyl groups on the piperidine ring brought about a dramatic change in the pharmacological profile of 2-[(cis- and trans-3,5-dimethylpiperidinyl)ethyl]-4-amino-5-chloro-2- methoxybenzoate, 7g,h. 7g (Ki = 0.26 +/- 0.06 nM) inhibited the relaxant action of 5-HT in the rat esophagus muscle with a pA2 value of 8.6. The advantage of the ester function was demonstrated by comparing the activity of several such compounds at 5-HT4 receptors with those of the corresponding amidic derivatives. This difference was less marked when the basic moiety was sterically constrained as in the quinuclidine and tropane moieties. Structural analyses of 7a,g were performed by determining their X-ray crystal structures and by molecular modeling (SYBYL). A relatively limited number of minimum energy conformers was found for both compounds. They were characterized by the cis folded conformation of the ethyl chain and by the orientation of the lone pair of the nitrogen atom pointing out of the molecule as seen in conformationally-constrained benzamides such as zacopride and renzapride. A hypothetical model for the 5-HT4 receptor with two sites for the binding of agonist and antagonist molecules was proposed.

New melatonin-like agents were designed from the frameworks of 2,5-dimethoxyphenethylamine, an important structural moiety for the 5-HT receptor, and (2-methoxynaphthyl)-ethylamine. The compounds were synthesized by classical methods and evaluated in binding assays with chicken brain membranes using 2-[125I]iodomelatonin as the radioligand. Preliminary studies on the series of N-acyl-disubstituted phenethylamines showed the favorable role of the methoxy group in the ortho position of the side chain on the affinity for the receptor (Ki = 8 +/- 0.2 nM) for N-[2-(2-methoxy-5-bromophenyl)ethyl]propionamide (3o). This effect was confirmed in a series of the naphthalene derivatives, a bioisosteric moiety of the indole ring, and several potent ligands for melatonin binding sites were prepared such as N-[2-(2-methoxynaphthyl)ethyl]propionamide (4b) (Ki = 0.67 +/- 0.05 nM) and N-[2-(2,7-dimethoxynaphthyl)ethyl]cyclopropylformamide (Ki = 0.05 +/- 0.004 nM) (4k). Structure-activity relationships are discussed with regard to melatonin and bioisosteric naphthalenic compound 2. The Ki value for 4b was affected to a similar extent to that of melatonin by GTP-gamma-S or Mn2+ in competition experiments, suggesting an agonist profile for this compound.

A three-dimensional quantitative structure-activity relationship using the comparative molecular field analysis (CoMFA) paradigm applied to 57 melatonin receptor ligands belonging to diverse structural families was performed. The compounds studied which have been synthesized previously and reported to be active at chicken brain melatonin receptors were divided into a training set of 48 molecules and a test set of 9 molecules. As most of these compounds have a highly flexible ethylamido side chain, the alignments were based on the most sterically constrained molecule which contains a tricyclic phenalene structure. This tricyclic compound can adopt an axial and an equatorial conformation. Two different molecular superpositions representing possible positioning within the receptor site have been suggested previously. CoMFA was tentatively used to discriminate between alternate hypothetical biologically active conformation and between possible positionings. The best 3D quantitative structure-activity relationship model found yields significant cross-validated, conventional, and predictive r2 values equal to 0.798, 0.967, and 0.76, respectively, along with an average absolute error of prediction of 0.25 log units. These results suggest that the active conformation of the most flexible molecules including melatonin is in a folded form if we consider the spatial position of the ethylamido side chain relative to the aromatic ring.