(1R, 3S)-(-)-trans-PAT: a novel full-efficacy serotonin 5-HT2C receptor agonist with 5-HT2A and 5-HT2B receptor inverse agonist/antagonist activity

Abstract

The serotonin 5-HT(2A), 5-HT(2B), and 5-HT(2C) G protein-coupled receptors signal primarily through G alpha(q) to activate phospholipase C (PLC) and formation of inositol phosphates (IP) and diacylglycerol. The human 5-HT(2C) receptor, expressed exclusively in the central nervous system, is involved in several physiological and psychological processes. Development of 5-HT(2C) agonists that do not also activate 5-HT(2A) or 5-HT(2B) receptors is challenging because transmembrane domain identity is about 75% among 5-HT(2) subtypes. This paper reports 5-HT(2) receptor affinity and function of (1R,3S)-(-)-trans-1-phenyl-3-dimethylamino-1,2,3,4-tetrahydronaphthalene (PAT), a small molecule that produces anorexia and weight-loss after peripheral administration to mice. (-)-Trans-PAT is a stereoselective full-efficacy agonist at human 5-HT(2C) receptors, plus, it is a 5-HT(2A)/5-HT(2B) inverse agonist and competitive antagonist. The K(i) of (-)-trans-PAT at 5-HT(2A), 5-HT(2B), and 5-HT(2C) receptors is 410, 1200, and 37 nM, respectively. Functional studies measured activation of PLC/[(3)H]-IP formation in clonal cells expressing human 5-HT(2) receptors. At 5-HT(2C) receptors, (-)-trans-PAT is an agonist (EC(50) = 20 nM) comparable to serotonin in potency and efficacy. At 5-HT(2A) and 5-HT(2B) receptors, (-)-trans-PAT is an inverse agonist (IC(50) = 490 and 1,000 nM, respectively) and competitive antagonist (K(B) = 460 and 1400 nM, respectively) of serotonin. Experimental results are interpreted in light of molecular modeling studies indicating the (-)-trans-PAT protonated amine can form an ionic bond with D3.32 of 5-HT(2A) and 5-HT(2C) receptors, but, not with 5-HT(2B) receptors. In addition to probing 5-HT(2) receptor structure and function, (-)-trans-PAT is a novel lead regarding 5-HT(2C) agonist/5-HT(2A) inverse agonist drug development for obesity and neuropsychiatric disorders.

Fig. 1

Structures of 1-phenyl-3-dimethylamino-1,2,3,4-tetrahydronaphthalene (PAT) stereoisomers.

Fig. 2

Representative saturation binding curves for serotonin 5-HT₂ receptor radioligands used to calculate B_max ± SEM. (pmol/mg protein) and K_d ± S.E.M (nM) values.

1. (A) [³H]-Ketanserin labeled 5-HT_2A receptors; B_max = 1.73 ± 0.11, K_d = 0.80 ± 0.03.

2. (B) [³H]-Mesulergine labeled 5-HT_2B receptors; B_max = 1.13 ± 0.39, K_d = 5.19 ± 0.36.

3. (C) [³H]-Mesulergine labeled 5-HT_2C receptors; B_max = 8.37 ± 0.15, K_d = 0.88 ± 0.03.

Fig. 3

Representative concentration–response curves for PAT isomer displacement of 5-HT₂ receptor radioligands from 5-HT_2A (A), 5-HT_2B (B), and 5-HT_2C (C) receptors. K_i and n_H values are summarized in the Table.

Fig. 4

Representative concentration–response curves for serotonin (closed squares) and (−)-trans-PAT (closed circles) activation of PLC/[³H]-IP formation in clonal cells expressing 5-HT_2C receptors. Single concentration effect of (−)-trans-PAT at 5-HT_2A (open triangles) and 5-HT_2B (open squares) receptors also is shown. EC₅₀ and n_H values are given in the text.

Fig. 5

Representative graphed data for (−)-trans-PAT competitive antagonism of serotonin activation of 5-HT_2A (A) and 5-HT_2B (B) receptors.

Fig. 6

Representative graphed data for (−)-trans-PAT inverse agonist activity at serotonin 5-HT_2A (A)and 5-HT_2B (B) receptors.

Fig. 7

Molecular models of the 5-HT_2A (A), 5-HT2B (B), and 5-HT_2C (C) receptors with (−)-trans-PAT docked at the putative binding pocket.

Fig. 7

Molecular models of the 5-HT_2A (A), 5-HT2B (B), and 5-HT_2C (C) receptors with (−)-trans-PAT docked at the putative binding pocket.

Fig. 7

Molecular models of the 5-HT_2A (A), 5-HT2B (B), and 5-HT_2C (C) receptors with (−)-trans-PAT docked at the putative binding pocket.