Highly Selective Dopamine D3 Receptor (D3R) Antagonists and Partial Agonists Based on Eticlopride and the D3R Crystal Structure: New Leads for Opioid Dependence Treatment

Abstract

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.

Figure 1

Representative D₃R antagonists

Figure 2

Design of the primary pharmacophore (PP)

Figure 3. Mouse Microsomal Phase I metabolism data for Compound 19

Compound 19 was incubated in mouse liver microsomes with NADPH regenerating system and compound disappearance was measured over time via LC/MS/MS. Compound 19 showed excellent metabolic stability substantiating its use in in vivo efficacy studies.

Figure 4

Effects of compound 19 (0, 5, 15 mg/kg, i.p.) on basal and oxycodone-induced increases in locomotor activity in mice. A: The effects of oxycodone on locomotion over time (from day 2 to day 9) in the presence or absence of compound 19 treatment. On day 1, compound 19 alone (5, 15 mg/kg, 15 min before saline) was administered and had no effect on locomotion. From day 2 to day 6, each animal received one daily dose of 19 (vehicle, 5, 15 mg/kg, 15 min prior to oxycodone) and oxycodone (4 mg/kg) administration. In the vehicle treatment group, repeated daily administration of oxycodone produced a progressive increase in locomotion (i.e., locomotor sensitization) (^#p<0.05, ^##p<0.01, compared to the level of locomotor activity on day 2). This locomotor sensitization was dose-dependently blocked by 19 (*p<0.05, **p<0.01, compared to the vehicle control group at each time point labeled). After 5 days of the (vehicle/19 + oxycodone) co-administration, animals underwent 2 days of withdrawal, followed by a 4 mg/kg oxycodone challenge injection (without 19 pretreatment) on day 9, indicating that oxycodone-induced locomotor sensitization was still present in the vehicle control group, which was blocked in the 19 pretreatment groups. B: The time course of 19-induced changes in locomotion (day 1), indicating that it has no effect on locomotor activity by itself. C: Time course of oxycodone-induced changes in locomotion after the first injection of oxycodone (acute effect on day 2), indicating that 19 pretreatment dose-dependently inhibited oxycodone-induced hyperactivity (acute effect). D: The time course of oxycodone priming-induced changes in locomotion in mice after 2 days of withdrawal from the last (vehicle/19 + oxycodone) co-administration (day 9), indicating that repeated 19 pretreatment from day 2 to day 6 produced a long-lasting inhibition in oxycodone-induced increases in locomotion. N=8 mice in each group.

Figure 5

Effects of compound 19 on oxycodone-induced conditioned place preference. Oxycodone (3 mg/kg, i.p.) conditioning in the (Vehicle + oxycodone) produced a robust place preference to the oxycodone-paired compartment. Pretreatment with 19 (5, 15 mg/kg, i.p., 15 min before oxycodone injection) dose-dependently blocked the acquisition of oxycodone-induced CPP. Compound 19 alone (15 mg/kg, i.p.) in the (19 + saline) group did not produce significant place preference. ** p<0.01, compared to the (Vehicle + vehicle) group; ^## p<0.01, compared to the (Vehicle + oxycodone) group. The number on each bar shows the animal number in each experimental group.

Scheme 1. Synthesis of analogues synthons 11–13 and full-length analogues 18–31^a

^aReagents and conditions: (a) fuming HNO₃, 0 °C to room temperature, 2h; (b) BH₃.CS₂, 24h; (c) PCC, CH₂Cl₂, overnight; (d) Ph₃P⁺CH₃Br^-, n-BuLi, THF; (e) 10% Pd/C, H₂, 50 psi, EtOH/EtOAc, 3h; (f) bis(2-chloroethyl)amine. HCl, diethyleneglycol monoethylether, 150 °C, 7h; (g) HCHO, NaBH(OAc)₃, AcOH; (h) 1-bromobutane, K₂CO₃, acetone, reflux, 7h; (i) n-bromobutyl phthalimide, K₂CO₃, acetone, reflux, 7h; (j) 2-(2-oxiran-2-yl)ethyl)isoindoline-1,3-dione, IPA, reflux, overnight; (k) hydrazine, EtOH, reflux, overnight; (l) ArCOOH, CDI, THF/DMF, 0 °C to room temperature; (m) ethyl 4-ethyl-1H-imidazole-2-carboxylate, (CH₃)₃Al, CH₂Cl₂, room temperature.

Scheme 2. Synthesis of 35^a

^aReagents and conditions: (a) 11b, K₂CO₃, acetone, reflux, overnight; (b) hydrazine, EtOH, reflux, overnight; (c) 4-methyl-1H-imidazole-2-carboxylic acid, CDI, THF, 0 °C to room temperature, overnight.

Scheme 3. Synthesis of 40^a

^aReagents and conditions: (a) 11b, K₂CO₃, acetone, reflux, overnight; (b) NaN₃, NH₄Cl, DMF, 100 °C, 6 h; (c) 10% Pd/C, H₂, 50 psi, 2 h; (d) Indole-2-COOH, CDI, THF, 0 °C to room temperature.