20 research outputs found
Molecular Determinants of the Intrinsic Efficacy of the Antipsychotic Aripiprazole
Partial agonists of the dopamine D2 receptor (D2R) have been developed to treat the symptoms of schizophrenia without causing the side effects elicited by antagonists. The receptor-ligand interactions that determine the intrinsic efficacy of such drugs, however, are poorly understood. Aripiprazole has an extended structure comprising a phenylpiperazine primary pharmacophore and a 1,2,3,4-tetrahydroquinolin-2-one secondary pharmacophore. We combined site-directed mutagenesis, analytical pharmacology, ligand fragments and molecular dynamics simulations to identify the D2R-aripiprazole interactions that contribute to affinity and efficacy. We reveal that an interaction between the secondary pharmacophore of aripiprazole and a secondary binding pocket defined by residues at the extracellular portions of transmembrane segments 1, 2 and 7 determine the intrinsic efficacy of aripiprazole. Our findings reveal a hitherto unappreciated mechanism through which to fine-tune the intrinsic efficacy of D2R agonists
Discovery of a novel class of negative allosteric modulator of the dopamine D2 receptor through fragmentation of a bitopic ligand
We recently demonstrated that SB269652 (1) engages one protomer of a dopamine D2 receptor (D2R) dimer in a bitopic mode to allosterically inhibit the binding of dopamine at the other protomer. Herein, we investigate structural deter- minants for allostery, focusing on modifications to three moieties within 1. We find that orthosteric âheadâ groups with small 7-substituents were important to maintain the limited negative cooperativity of analogues of 1, and replacement of the tetrahydroisoquinoline head group with other D2R âprivileged structuresâ generated orthosteric antagonists. Additionally, replacement of the cyclohexylene linker with polymethylene chains conferred linker length dependency in allosteric pharmacology. We validated the importance of the indolic NH as a hydrogen bond donor moiety for maintaining allostery. Replacement of the indole ring with azaindole conferred a 30-fold increase in affinity while maintaining negative cooperativity. Combined, these results provide novel SAR insight for bitopic ligands that act as negative allosteric modulators of the D2R
Understanding biased agonism at the dopamine Dâ receptor
The phenomenon of âbiased agonismâ presents an attractive avenue for drug development as it allows the separation of therapeutic effects from side effects mediated by the same target. A prototypical G protein-coupled receptor at which biased agonism has been extensively studied is the dopamine Dâ receptor, an important therapeutic target for current treatments of Parkinsonâs disease and schizophrenia. There is increasing evidence that biased agonism is important for the antipsychotic efficacy of dopamine Dâ receptor partial agonists, such as aripiprazole and cariprazine. However, a clear relationship between biased agonism at the dopamine Dâ receptor and antipsychotic efficacy remains elusive, not least due to discrepancies in literature describing aripiprazole as âbiasedâ or âunbiasedâ, despite the same signalling endpoints being studied using the same cell background. To clarify such conflicts, and to aid the drug discovery efforts aimed at identifying novel dopamine Dâ receptor biased agonists, the focus of this thesis is to gain greater insight into the mechanisms that mediate biased agonism at the dopamine Dâ receptor. Through the utilization of both mutagenesis-based and structure-activity-based approaches, a secondary binding pocket was identified for being crucial in the affinity, efficacy, and bias of different ligands at the dopamine Dâ receptor. A structure-activity relationship study indicated that both efficacy and biased agonism can be finely tuned by minor structural modifications to the head group, the tail group, and the orientation and length of a spacer region of cariprazine. In particular, it was demonstrated that modifications to the tail region, and thus the interaction with a potential secondary binding site, alter the orientation of the head group within the orthosteric binding site regulating both efficacy and biased agonism. These results were corroborated with a mutagenesis study, in which mutations within a putative secondary binding site significantly impacted the affinity and efficacy of a number of dopamine Dâ receptor agonists. Finally, it was demonstrated that âkinetic contextâ, as determined by both ligand-binding kinetics and the kinetics intrinsic to different cellular signalling processes, can dramatically impact observations of biased agonism. Such findings illustrate, for the first time, the importance of incorporating kinetic profiling in future studies focussed on biased agonism to allow a more informed selection of preclinical candidates and thus an improved foundation for drug discovery of biased agonists
StructureâActivity Relationships of Privileged Structures Lead to the Discovery of Novel Biased Ligands at the Dopamine D 2Receptor
Biased agonism at GPCRs highlights the potential for the discovery and design of pathway-selective ligands and may confer therapeutic advantages to ligands targeting the dopamine D2 receptor (D2R). We investigated the determinants of efficacy, affinity, and bias for three privileged structures for the D2R, exploring changes to linker length and incorporation of a heterocyclic unit. Profiling the compounds in two signaling assays (cAMP and pERK1/2) allowed us to identify and quantify determinants of biased agonism at the D2R. Substitution on the phenylpiperazine privileged structures (2-methoxy vs 2,3-dichloro) influenced bias when the thienopyridine heterocycle was absent. Upon inclusion of the thienopyridine unit, the substitution pattern (4,6-dimethyl vs 5-chloro-6-methoxy-4-methyl) had a significant effect on bias that overruled the effect of the phenylpiperazine substitution pattern. This latter observation could be reconciled with an extended binding mode for these compounds, whereby the interaction of the heterocycle with a secondary binding pocket may engender bias
Proof of concept study for designed multiple ligands targeting the dopamine D2, serotonin 5-HT2A, and muscarinic M1 acetylcholine receptors
Herein we describe the hybridization of a benzoxazinone M1 scaffold with D2 privileged structures derived from putative and clinically relevant antipsychotics to develop designed multiple ligands. The M1 mAChR is an attractive target for the cognitive deficits in key CNS disorders. Moreover, activity at D2 and 5-HT2A receptors has proven useful for antipsychotic efficacy. We identified 9 which retained functional activity at the target M1 mAChR and D2R and demonstrated high affinity for the 5-HT2AR
StructureâActivity Relationships of Privileged Structures Lead to the Discovery of Novel Biased Ligands at the Dopamine D<sub>2</sub> Receptor
Biased agonism at GPCRs highlights
the potential for the discovery
and design of pathway-selective ligands and may confer therapeutic
advantages to ligands targeting the dopamine D<sub>2</sub> receptor
(D<sub>2</sub>R). We investigated the determinants of efficacy, affinity,
and bias for three privileged structures for the D<sub>2</sub>R, exploring
changes to linker length and incorporation of a heterocyclic unit.
Profiling the compounds in two signaling assays (cAMP and pERK1/2)
allowed us to identify and quantify determinants of biased agonism
at the D<sub>2</sub>R. Substitution on the phenylpiperazine privileged
structures (2-methoxy vs 2,3-dichloro) influenced bias when the thienopyridine
heterocycle was absent. Upon inclusion of the thienopyridine unit,
the substitution pattern (4,6-dimethyl vs 5-chloro-6-methoxy-4-methyl)
had a significant effect on bias that overruled the effect of the
phenylpiperazine substitution pattern. This latter observation could
be reconciled with an extended binding mode for these compounds, whereby
the interaction of the heterocycle with a secondary binding pocket
may engender bias
A structure-activity analysis of biased agonism at the dopamine D2 receptor.
Biased agonism offers an opportunity for the medicinal chemist to discover pathway-selective ligands for GPCRs. A number of studies have suggested that biased agonism at the dopamine D2 receptor (D2R) may be advantageous for the treatment of neuropsychiatric disorders, including schizophrenia. As such, it is of great importance to gain insight into the SAR of biased agonism at this receptor. We have generated SAR based on a novel D2R partial agonist, tert-butyl (trans-4-(2-(3,4-dihydroisoquinolin-2(1H)-yl)ethyl)cyclohexyl)carbamate (4). This ligand shares structural similarity to cariprazine (2), a drug awaiting FDA approval for the treatment of schizophrenia, yet displays a distinct bias toward two different signaling end points. We synthesized a number of derivatives of 4 with subtle structural modifications, including incorporation of cariprazine fragments. By combining pharmacological profiling with analytical methodology to identify and to quantify bias, we have demonstrated that efficacy and biased agonism can be finely tuned by minor structural modifications to the head group containing the tertiary amine, a tail group that extends away from this moiety, and the orientation and length of a spacer region between these two moieties
A Novel Nonribose Agonist, LUF5834, Engages Residues That Are Distinct from Those of Adenosine-Like Ligands to Activate the Adenosine A2a Receptor
The recent publication of both the antagonist- and agonist-bound structures of the adenosine A(2A) receptor have revealed much about how a ligand may bind to a receptor and cause the conformational changes associated with agonist-mediated activation. In particular, the agonist-bound structure revealed key interactions between the ribose group of adenosine-derived agonists and amino acids in the receptor binding pocket that lead to receptor activation. However, agonists without a ribose group also exist, and we wondered whether such compounds occupy the same agonist binding site. Therefore we used a mutagenesis approach in this study to investigate the mode of binding of 2-amino-4-(4-hydroxyphenyl)- 6-(1H-imidazol-2-ylmethylsulfanyl)pyridine-3,5-dicarbonitrile (LUF5834), a potent partial agonist without a ribose moiety, compared with the adenosine-derived reference agonist 2-[p-(2-carboxyethyl)phenyl-ethylamino]-5'-N-ethylcarboxamidoadenosine (CGS21680). Mutation of the orthosteric residue Phe168 to alanine abrogated the function of both agonists. However, mutation to alanine of residues Thr88 and Ser277 shown by the crystal structures to interact with the ribose group of adenosine-like ligands had no effect on the potency of LUF5834. Furthermore, alanine mutation of Asn253, which makes a hydrogen-bonding interaction with the exocyclic nitrogen of the adenine ring, had minimal effect on LUF5834 affinity but removed agonist activity of this ligand. Mutation of other residues, such as the highly conserved Trp246 or Glu13, had significant deleterious effects on the function of CGS21680 but little effect on LUF5834. In summary, our findings suggest that this class of agonist interacts with distinct residues to activate the receptor compared with classic adenosine derived agonists
Proof of Concept Study for Designed Multiple Ligands Targeting the Dopamine D<sub>2</sub>, Serotonin 5âHT<sub>2A</sub>, and Muscarinic M<sub>1</sub> Acetylcholine Receptors
Herein we describe the hybridization
of a benzoxazinone M<sub>1</sub> scaffold with D<sub>2</sub> privileged
structures derived from putative
and clinically relevant antipsychotics to develop designed multiple
ligands. The M<sub>1</sub> mAChR is an attractive target for the cognitive
deficits in key CNS disorders. Moreover, activity at D<sub>2</sub> and 5-HT<sub>2A</sub> receptors has proven useful for antipsychotic
efficacy. We identified <b>9</b> which retained functional activity
at the target M<sub>1</sub> mAChR and D<sub>2</sub>R and demonstrated
high affinity for the 5-HT<sub>2A</sub>R