Design of a True Bivalent Ligand with Picomolar Binding Affinity for a G Protein-Coupled Receptor Homodimer

Abstract

Bivalent ligands have emerged as chemical tools to study G protein-coupled receptor dimers. Using a combination of computational, chemical, and biochemical tools, here we describe the design of bivalent ligand 13 with high affinity (K DB1 = 21 pM) for the dopamine D 2 receptor (D 2 R) homodimer. Bivalent ligand 13 enhances the binding affinity relative to monovalent compound 15 by 37-fold, indicating simultaneous binding at both protomers. Using synthetic peptides with amino acid sequences of transmembrane (TM) domains of D 2 R, we provide evidence that TM6 forms the interface of the homodimer. Notably, the disturber peptide TAT-TM6 decreased the binding of bivalent ligand 13 by 52-fold and had no effect on monovalent compound 15, confirming the D 2 R homodimer through TM6 ex vivo. In conclusion, by using a versatile multivalent chemical platform, we have developed a precise strategy to generate a true bivalent ligand that simultaneously targets both orthosteric sites of the D 2 R homodimer.

Distinct Dopamine D2 Receptor Antagonists Differentially Impact D2 Receptor Oligomerization

“…In order to probe how a D 2 R homodimer might be affected by sidechain conformational changes in TM5 and TM6, a D 2 R homodimer without bound antagonist was modeled from the original crystal structure with a TM5–TM6–TM5–TM6 interface (in line with experimental evidence by Pulido, 2018 [32]) by protein–protein docking. This resulted in a D 2 R homodimer with a highly favorable interface docking score of −9.7 (on a scale of 0 to −10, where lower than −5.0 was considered satisfactory ([58]); see Methods).…”

“…Different transmembrane (TM) regions of the D 2 R have been reported to be involved in the D 2 R homodimer interface. Incubation of D 2 R homodimers with peptides derived from the putative TM6 regions of the D 2 R resulted in dissociation of the dimer to the monomers [24,32]. On the other hand, successive deletion of TM domains of the D 2 R and cysteine cross-linking studies revealed that the most critical areas involved in the intermolecular hydrophobic interactions for dimerization resided in TM4 [33,34].…”

Dopamine Receptor Subtypes, Physiology and Pharmacology: New Ligands and Concepts in Schizophrenia

“…It has allowed to show the presence of an alternative mechanism of D3 receptor internalization independent of β-arrestin and used by group II GPCR ( Xu et al., 2019 ). Considering the excess D2 homodimers detected in schizophrenia ( Wang et al., 2010 ), the effects of DA antagonists on these entities has been specifically explored using bivalent ligands ( Pulido et al., 2018 ; Wouters et al., 2019 ). A molecular model of the homodimer has been also generated for D2 to provide docking information relative to bivalent ligands with different pharmacological properties (for example orthosteric and allosteric agents) ( Kaczor et al., 2016 ).…”

The Endocannabinoid System as a Target in Cancer Diseases: Are We There Yet?

“…Thus, the receptor heteromer would be more likely to be disease-specific than would the corresponding monomeric/homomeric receptors” (Cortés et al, 2016). Newer reports show that heteromer-selective drugs can exist in the form of small-molecules, bivalent or multifunctional ligands, or antibodies, displaying higher affinity and efficacy for a receptor that forms a certain heteromer than for this receptor in another heteromer or in the monomeric form (Orru et al, 2011; Gomes et al, 2013c,b, 2016a; Cortés et al, 2016; Pulido et al, 2018; Qian et al, 2018a,b).…”