Ligand discovery from a dopamine D3 receptor homology model and crystal structure

Abstract

G protein-coupled receptors (GPCRs) are intensely studied as drug targets and for their role in signaling. With the determination of the first crystal structures, interest in structure-based ligand discovery increased. Unfortunately, for most GPCRs no experimental structures are available. The determination of the D(3) receptor structure and the challenge to the community to predict it enabled a fully prospective comparison of ligand discovery from a modeled structure versus that of the subsequently released crystal structure. Over 3.3 million molecules were docked against a homology model, and 26 of the highest ranking were tested for binding. Six had affinities ranging from 0.2 to 3.1 μM. Subsequently, the crystal structure was released and the docking screen repeated. Of the 25 compounds selected, five had affinities ranging from 0.3 to 3.0 μM. One of the new ligands from the homology model screen was optimized for affinity to 81 nM. The feasibility of docking screens against modeled GPCRs more generally is considered.

Figure 1. Predicted Structure of the Dopamine D₃ Receptor Binding Site

(a) Comparison of the homology model of the dopamine D₃ receptor in complex with eticlopride (light blue) to the crystal structure (yellow) visualized with PyMOL. The structures have been aligned using 15 binding site residues. Polar interactions for the crystal structure are shown in black dotted lines. (b) Chemical structure of eticlopride (compound 1).

Figure 1. Predicted Structure of the Dopamine D₃ Receptor Binding Site

(a) Comparison of the homology model of the dopamine D₃ receptor in complex with eticlopride (light blue) to the crystal structure (yellow) visualized with PyMOL. The structures have been aligned using 15 binding site residues. Polar interactions for the crystal structure are shown in black dotted lines. (b) Chemical structure of eticlopride (compound 1).

Figure 2. Predicted binding modes of ligands found from the homology model screen

Predicted binding poses for four ligands discovered in the docking screen against the dopamine D₃ receptor homology model, visualized with UCSF Chimera: (a) 3 (b) 4 (c) 6 (d) 7 Predicted binding modes for the two analogs of compound 3 based on docking to the homology model (e) 56 and (f) 57.

Figure 3. Dose-response curves of discovered ligands

Representative radioligand ([³H]N-methylspiperone) competition binding isotherms for compounds 3, 4, 7, 28, 30 and 31 (a-f). Data for a reference compound (chlorpromazine, black curve) are shown along with data for the test compound (red curve). Assays are performed using a final radioligand concentration between (0.5 × K_D) and (1 × K_D), where K_D equals the radioligand dissociation constant, which is determined for each crude membrane preparation by radioligand saturation binding analysis. Data represent mean values ± standard error, performed on triplicate experiments.

Figure 4. Predicted binding modes of ligands found from the crystal structure screen

Predicted binding poses for the ligands discovered in the docking screen against the dopamine D₃ receptor crystal structure, visualized with UCSF Chimera: (a) 28 (b) 29 (c) 31 (d) 32.