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The dopamine D4 receptor (D4R), a G protein-coupled receptor, is predominantly expressed in brain regions that control cognition, attention, and decision making. Previous studies have indicated that D4R-targeted ligands could be promising therapeutic targets for the treatment of several neuropsychiatric conditions, including substance use disorders (SUDs). There are currently no FDA-approved medications that selectively target D4Rs. New ligands may facilitate better understanding of the role of D4R-mediated signaling in drug-taking and drug-seeking behaviors. The present study focuses on the synthesis and evaluation of a novel series of benzothiazole analogues designed to target D4R. We identified several compounds with high D4R binding affinity (Ki ≤ 6.87 nM) and >91-fold selectivity over other D2-like receptors (D2R, D3R) with diverse partial agonist and antagonist profiles. Based on receptor affinity and functional analyses, 5f was identified as a potent low-efficacy partial agonist of the D4R and selected for further investigation. 5f was metabolically stable in rat and human liver microsome assays and displayed excellent brain penetration in rats. Using a within-session multidosing procedure, 5f (5, 15 and 30 mg/kg, i.p.) dose-dependently decreased i.v. infusions of three-unit doses of cocaine under a fixed-ratio (FR) FR3 schedule of reinforcement. These results are consistent with previous results produced by D4R-selective antagonists in SUD models, however off-target antagonism of 5-HT2A or 5-HT2B receptors may also contribute to these effects. Results with compound 5f support further efforts to target D4R in the treatment of SUDs. Further development of the benzothiazole scaffold may engineer out any serotonergic activity.

Linkers are emerging as a key component in regulating the pharmacology of bitopic ligands directed toward G-protein coupled receptors (GPCRs). In this study, the role of regio- and stereochemistry in cyclic aliphatic linkers tethering well-characterized primary and secondary pharmacophores targeting dopamine D2 and D3 receptor subtypes (D2R and D3R, respectively) is described. We introduce several potent and selective D2R (rel-trans-16b; D2R Ki = 4.58 nM) and D3R (rel-cis-14a; D3R Ki = 5.72 nM) agonists while modulating subtype selectivity in a stereospecific fashion, transferring D2R selectivity toward D3R via inversion of the stereochemistry around these cyclic aliphatic linkers [e.g., (-)-(1S,2R)-43 and (+)-(1R,2S)-42]. Pharmacological observations were supported with extensive molecular docking studies. Thus, not only is it an innovative approach to modulate the pharmacology of dopaminergic ligands described, but a new class of optically active cyclic linkers are also introduced, which can be used to expand the bitopic drug design approach toward other GPCRs.

The crystal structure of the dopamine D3 receptor (D3R) in complex with eticlopride inspired the design of bitopic ligands that explored (1) N-alkylation of the eticlopride's pyrrolidine ring, (2) shifting of the position of the pyrrolidine nitrogen, (3) expansion of the pyrrolidine ring system, and (4) incorporation of O-alkylations at the 4-position. Structure activity relationships (SAR) revealed that moving the N- or expanding the pyrrolidine ring was detrimental to D2R/D3R binding affinities. Small pyrrolidine N-alkyl groups were poorly tolerated, but the addition of a linker and secondary pharmacophore (SP) improved affinities. Moreover, O-alkylated analogues showed higher binding affinities compared to analogously N-alkylated compounds, e.g., O-alkylated 33 (D3R, 0.436 nM and D2R, 1.77 nM) vs the N-alkylated 11 (D3R, 6.97 nM and D2R, 25.3 nM). All lead molecules were functional D2R/D3R antagonists. Molecular models confirmed that 4-position modifications would be well-tolerated for future D2R/D3R bioconjugate tools that require long linkers and or sterically bulky groups.

In the search for novel bitopic compounds targeting the dopamine D3 receptor (D3R), the N-(2,3-dichlorophenyl)piperazine nucleus (primary pharmacophore) has been linked to the 6,6- or 5,5-diphenyl-1,4-dioxane-2-carboxamide or the 1,4-benzodioxane-2-carboxamide scaffold (secondary pharmacophore) by an unsubstituted or 3-F-/3-OH-substituted butyl chain. This scaffold hybridization strategy led to the discovery of potent D3R-selective or multitarget ligands potentially useful for central nervous system disorders. In particular, the 6,6-diphenyl-1,4-dioxane derivative 3 showed a D3R-preferential profile, while an interesting multitarget behavior has been highlighted for the 5,5-diphenyl-1,4-dioxane and 1,4-benzodioxane derivatives 6 and 9, respectively, which displayed potent D2R antagonism, 5-HT1AR and D4R agonism, as well as potent D3R partial agonism. They also behaved as low-potency 5-HT2AR antagonists and 5-HT2CR partial agonists. Such a profile might be a promising starting point for the discovery of novel antipsychotic agents.

The dopamine D4 receptor (D4R), a G protein-coupled receptor, is predominantly expressed in the prefrontal cortex in which it plays an important role in cognition, attention, and decision making. Studies have indicated D4R-selective ligands as promising therapeutic targets for the treatment of neuropsychiatric conditions such as substance use disorders (SUD). D4R ligands have been shown to alter cognition and behavior in animal models of drug addiction. A better understanding of D4R-mediated signaling is essential to treating D4R-associated disorders, including SUD. Despite its clinical importance, there are currently no FDA approved medications that target the D4R, and for example, treat cocaine use disorder. The present study focuses on the design of D4R-selective ligands based on the parental phenylpiperazine scaffold and pharmacokinetic analysis in rat and human liver microsomes, followed by in vivo pharmacokinetic and behavioral analysis. We identified several compounds with high binding affinity and D4R selectivity (Ki ≤ 6.87 nM and >91-fold vs. other D2-like receptors (D2R, D3R)) with diverse partial agonist and antagonist profiles. Based on the affinity profiles, 5f (CAB-01-019) was selected for an in vitro metabolic stability assay in rat and human liver microsomes which displayed an acceptable stability profile. Based on these results, we moved 5f forward into in vivo pharmacokinetic analysis in rats where it displayed excellent brain penetration with AUCbrain/plasma > 3. The full antagonist 5f (5, 15 and 30 mg/kg, i.p.) was tested in cocaine self-administration studies in rats, using a within-session multidosing procedure. 5f dose-dependently decreased the number of i.v. infusions obtained for three-unit doses of cocaine under a fixed ratio (FR) FR3 schedule of reinforcement, suggesting that the selected D4R antagonist reduced the rewarding effects of cocaine. Together, these results support the development of D4R-selective antagonists for SUDs and support 5f as a new probe for studying D4R-specific behaviors.

The need for safer pain-management therapies with decreased abuse liability inspired a novel drug design that retains μ-opioid receptor (MOR)-mediated analgesia, while minimizing addictive liability. We recently demonstrated that targeting the dopamine D3 receptor (D3R) with highly selective antagonists/partial agonists can reduce opioid self-administration and reinstatement to drug seeking in rodent models without diminishing antinociceptive effects. The identification of the D3R as a target for the treatment of opioid use disorders prompted the idea of generating a class of ligands presenting bitopic or bivalent structures, allowing the dual-target binding of the MOR and D3R. Structure-activity relationship studies using computationally aided drug design and in vitro binding assays led to the identification of potent dual-target leads (23, 28, and 40), based on different structural templates and scaffolds, with moderate (sub-micromolar) to high (low nanomolar/sub-nanomolar) binding affinities. Bioluminescence resonance energy transfer-based functional studies revealed MOR agonist-D3R antagonist/partial agonist efficacies that suggest potential for maintaining analgesia with reduced opioid-abuse liability.

In this paper, the benzo-cracking approach was applied to the potent sigma1 (σ1) receptor antagonist 1 to afford the less conformationally constrained 1,3-dioxane derivatives 2 and 3. To evaluate the effect of the increase in the distance between the two hydrophobic structural elements that flank the basic function, the cis and trans diastereomers of 4 and 5 were also prepared and studied. Compounds 2 and 3 showed affinity values at the σ1 receptor significantly higher than that of the lead compound 1. In particular, 3 displayed unprecedented selectivity over the σ2 receptor, the phencyclidine site of the NMDA receptor, and opioid receptor subtypes, as well as over the dopamine transporter. Docking results supported the structure–activity relationship studies. Due to its interesting biological profile, derivative 3, selected for an in vivo study in a validated preclinical model of binge eating, was able to counteract the overeating of palatable food only in binging rats, without affecting palatable food intake in the control group and anxiety-like and depression-related behaviors in female rats. This result strengthened the involvement of the σ1 receptor in the compulsive-like eating behavior and supported the σ1 receptor as a promising target for the management of eating disorders.

Missense mutations that give rise to protein misfolding are rare, but collectively, defective protein folding diseases are consequential. Folding deficiencies are amenable to pharmacological correction (pharmacochaperoning), but the underlying mechanisms remain enigmatic. Ibogaine and its active metabolite noribogaine correct folding defects in the dopamine transporter (DAT), but they rescue only a very limited number of folding-deficient DAT mutant proteins, which give rise to infantile Parkinsonism and dystonia. Herein, a series of analogs was generated by reconfiguring the complex ibogaine ring system and exploring the structural requirements for binding to wild-type transporters, as well as for rescuing two equivalent synthetic folding-deficient mutants, SERT-PG601,602AA and DAT-PG584,585AA. The most active tropane-based analog (9b) was also an effective pharmacochaperone in vivo in Drosophila harboring the DAT-PG584,585AA mutation and rescued 6 out of 13 disease-associated human DAT mutant proteins in vitro. Hence, a novel lead pharmacochaperone has been identified that demonstrates medication development potential for patients harboring DAT mutations.

A series of novel 1,4-dioxane analogues of the muscarinic acetylcholine receptor (mAChR) antagonist 2 was synthesized and studied for their affinity at M1–M5 mAChRs. The 6-cyclohexyl-6-phenyl derivative 3b, with a cis configuration between the CH2N+(CH3)3 chain in the 2-position and the cyclohexyl moiety in the 6-position, showed pKi values for mAChRs higher than those of 2 and a selectivity profile analogous to that of the clinically approved drug oxybutynin. The study of the enantiomers of 3b and the corresponding tertiary amine 33b revealed that the eutomers are (2S,6S)-(−)-3b and (2S,6S)-(−)-33b, respectively. Docking simulations on the M3 mAChR-resolved structure rationalized the experimental observations. The quaternary ammonium function, which should prevent the crossing of the blood–brain barrier, and the high M3/M2 selectivity, which might limit cardiovascular side effects, make 3b a valuable starting point for the design of novel antagonists potentially useful in peripheral diseases in which M3 receptors are involved.

In this study, starting from our selective D3R agonist FOB02-04A (5), we investigated the chemical space around the linker portion of the molecule via insertion of a hydroxyl substituent and ring-e...

Because of the large degree of homology among dopamine D2-like receptors, discovering ligands capable of discriminating between the D2, D3, and D4 receptor subtypes remains a significant challenge....

The discovery of functionally biased and physiologically beneficial ligands directed toward G-protein coupled receptors (GPCRs) has provided the impetus to design dopamine D2 receptor (D2R) targeted molecules that may be therapeutically advantageous for the treatment of certain neuropsychiatric or basal ganglia related disorders. Here we describe the synthesis of a novel series of D2R agonists linking the D2R unbiased agonist sumanirole with privileged secondary molecular fragments. The resulting ligands demonstrate improved D2R affinity and selectivity over sumanirole. Extensive in vitro functional studies and bias factor analysis led to the identification of a novel class of highly potent Go-protein biased full D2R agonists with more than 10-fold and 1000-fold bias selectivity toward activation of specific G-protein subtypes and β-arrestin, respectively. Intracellular electrophysiological recordings from midbrain dopamine neurons demonstrated that Go-protein selective agonists can elicit prolonged ligand-induced GIRK activity via D2Rs, which may be beneficial in the treatment of dyskinesias associated with dopamine system dysfunction.

Both dopamine D3 receptor (D3R) partial agonists and antagonists have been implicated as potential medications for substance use disorders. In contrast to antagonists, partial agonists may cause fewer side effects since they maintain some dopaminergic tone and may be less disruptive to normal neuronal functions. Here, we report three sets of 4-phenylpiperazine stereoisomers that differ considerably in efficacy: the (R)-enantiomers are antagonists/weak partial agonists, whereas the (S)-enantiomers are much more efficacious. To investigate the structural basis of partial agonism, we performed comparative microsecond-scale molecular dynamics simulations starting from the inactive state of D3R in complex with these enantiomers. Analysis of the simulation results reveals common structural rearrangements near the ligand binding site induced by the bound (S)-enantiomers, but not by the (R)-enantiomers, that are features of partially activated receptor conformations. These receptor models bound with partial agoni...

The recent and precipitous increase in opioid analgesic abuse and overdose has inspired investigation of the dopamine D3 receptor (D3R) as a target for therapeutic intervention. Metabolic instability or predicted toxicity has precluded successful translation of previously reported D3R-selective antagonists to clinical use for cocaine abuse. Herein, we report a series of novel and D3R crystal structure-guided 4-phenylpiperazines with exceptionally high D3R affinities and/or selectivities with varying efficacies. Lead compound 19 was selected based on its in vitro profile: D3R Ki = 6.84 nM, 1700 fold D3R versus D2R binding selectivity, and its metabolic stability in mouse microsomes. Compound 19 inhibited oxycodone-induced hyperlocomotion in mice and reduced oxycodone-induced locomotor sensitization. In addition, pretreatment with 19 also dose-dependently inhibited the acquisition of oxycodone-induced conditioned place preference (CPP) in rats. These findings support the D3R as a target for opioid dependence treatment and compound 19 as a new lead molecule for development.

Two series of 1,4-dioxanes (4-11 and 12-19) were rationally designed and prepared to interact either with the phencyclidine (PCP) binding site of the N-methyl-d-aspartate (NMDA) receptor or with σ1 receptors, respectively. The biological profiles of the novel compounds were assessed using radioligand binding assays, and the compounds with the highest affinities were investigated for their functional activity. The results were in line with the available pharmacophore models and highlighted that the 1,4-dioxane scaffold is compatible with potent antagonist activity at NMDA receptor or high affinity for σ1 receptors. The primary amines 6b and 7 bearing a cyclohexyl and a phenyl ring or two phenyl rings in position 6, respectively, were the most potent noncompetitive antagonists at the NMDA receptor with IC50 values similar to those of the dissociative anesthetic (S)-(+)-ketamine. The 5,5-diphenyl substitution associated with a benzylaminomethyl moiety in position 2, as in 18, favored the interaction with σ1 receptors.

5-HT1A receptor and α1-adrenoreceptor (α1-AR) binding sites recognized by the 1,4-dioxanes 2–4 display reversed stereochemical requirements. (S)-2 proved to be a potent 5-HT1A receptor agonist high...