Discovery of Guanfacine as a Novel TAAR1 Agonist: A Combination Strategy through Molecular Modeling Studies and Biological Assays

Abstract

Trace amine-associated receptor 1 (TAAR1) is an attractive target for the design of innovative drugs to be applied in diverse pharmacological settings. Due to a non-negligible structural similarity with endogenous ligands, most of the agonists developed so far resulted in being affected by a low selectivity for TAAR1 with respect to other monoaminergic G protein-coupled receptors, like the adrenoreceptors. This study utilized comparative molecular docking studies and quantitative-structure activity relationship (QSAR) analyses to unveil key structural differences between TAAR1 and alpha2-adrenoreceptor (α₂-ADR), with the aim to design novel TAAR1 agonists characterized by a higher selectivity profile and reduced off-target effects. While the presence of hydrophobic motives is encouraged towards both the two receptors, the introduction of polar/positively charged groups and the ligand conformation deeply affect the TAAR1 or α₂-ADR putative selectivity. These computational methods allowed the identification of the α₂A-ADR agonist guanfacine as an attractive TAAR1-targeting lead compound, demonstrating nanomolar activity in vitro. In vivo exploration of the efficacy of guanfacine showed that it is able to decrease the locomotor activity of dopamine transporter knockout (DAT-KO) rats. Therefore, guanfacine can be considered as an interesting template molecule worthy of structural optimization. The dual activity of guanfacine on both α₂-ADR and TAAR1 signaling and the related crosstalk between the two pathways will deserve more in-depth investigation.

Keywords: AlphaFold; GPCR; QSAR; TAAR1; docking; dopamine; guanfacine; α2-adrenoreceptor.

Figure 1

Chemical structure and biological activity of the reference dual acting TAAR1 and α₂-ADR ligand S18616. Representative examples of TAAR1 or α₂-ADR selective imidazoline and imidazole derivatives [35].

Figure 2

Workflow of the present study: a combination approach through molecular modeling studies and biological assays.

Figure 3

Docking mode of S18616 (C atom; magenta) at the hTAAR1 (A,B) and α₂-ADR (C,D) binding site. A perspective of the ligand volume (light brown) and of the protein cavity is depicted in (A) and (C), respectively. The most important residues involved in the agonist binding are labelled (B,D). A and C representations have been performed via Chimera 1.16 [51], while the ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 4

Docking pose of compound 5a (C atom; magenta) within the hTAAR1 (A) and α₂-ADR (B) binding sites. The most important residues involved in the agonist binding are labelled. Ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 5

Docking pose of compound 6a (C atom; magenta) within the hTAAR1 (A) and α₂-ADR (B) binding sites. The most important residues involved in the agonist binding are labelled. Ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 6

Docking pose of compound 5c (C atom; magenta) within the hTAAR1 (A) and α₂-ADR (B) binding sites. The most important residues involved in the agonist binding are labelled. Ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 7

Docking mode of 6c (C atom; magenta) at the hTAAR1 (A) and at the α₂-ADR (B) binding site. The most important residues involved in the agonist binding are labelled. Ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 8

Docking positioning of 16 (C atom; magenta) at the hTAAR1 binding site (A) and of 18 at the α₂-ADR cavity (B). The most important residues involved in the agonist binding are labelled. Ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 9

Docking mode of 24 (C atom; magenta) at the hTAAR1 (A) and at the α₂-ADR (B) binding site. The most important residues involved in the agonist binding are labelled. Ligand-protein contacts have been explored by means of PyMol software 2.5.2—Incentive Product Copyright (C) Schrodinger, LLC [52].

Figure 10

Distribution of the predicted (Pred. hTAAR1 pKi) versus the experimental (Exp. hTAAR1 pKi) hTAAR1 binding affinity featured by the training set (A) and test set derivatives (B). Compounds are represented as dots.

Figure 11

Distribution of the predicted (Pred. α₂-ADR pKi) versus the experimental (Exp. α₂-ADR pKi) α₂-ADR binding affinity featured by the training set (A) and test set derivatives (B). Compounds are represented as dots.

Figure 12

Schematic representation of the importance played by the DipoleY and Q_VSA_PNEG descriptors to influence the compound (hTAAR1 and α₂-ADR) binding affinity values. The two descriptors are endowed by the highest RI values in model (A,B), respectively. Compounds are represented as dots.

Figure 13

PM_A (hTAAR1 targeting ability) (A) and PM_B (α₂-ADR targeting ability) (B) as featured by the dataset compounds. The most relevant features (F) are represented as colored spheres and classified as related to aromatic or hydrophobic groups (AroǀHyd) or only hydrophobic- (Hyd) or aromatic- cores (Aro), or to H-bonding acceptor (Acc) or donor (Don) groups. Compounds 25 (C atom; light pink) and 14 (C atom; light pink) were taken as representative of the dataset in PM_A and PM_B, respectively. Distance among the recurrent features shared by both PM_A and PM_B are reported.

Figure 14

The functional activity at hTAAR1 of guanfacine, guanabenz, and β-PEA presented as concentration-dependent curves.

Figure 15

Action of guanfacine (0.1 and 0.3 mg/kg, i/p) on locomotor activity of WT and DAT-KO rats. ** p < 0.01, * p < 0.05 (Bonferroni’s test) vs. the vehicle-treated animals.