An improved synthesis of a haptenic heroin surrogate 1 (6-AmHap) is reported. The intermediate needed for the preparation of 1 was described in the route in the synthesis of 2 (DiAmHap). A scalable procedure was developed to install the C-3 amido group. Using the Boc protectng group in 18 allowed preparation of 1 in an overall yield of 53% from 4 and eliminated the necessity of preparing the diamide 13. Hapten 1 was conjugated to tetanus toxoid and mixed with liposomes containing monophosphoryl lipid A as an adjuvant. The 1 vaccine induced high anti-1 IgG levels that reduced heroin-induced antinociception and locomotive behavioral changes following repeated subcutaneous and intravenous heroin challenges in mice and rats. Vaccinated mice had reduced heroin-induced hyperlocomotion following a 50 mg/kg heroin challenge. The 1 vaccine-induced antibodies bound to heroin and other abused opioids, including hydrocodone, oxycodone, hydromorphone, oxymorphone, and codeine.

Synthetic cathinones are common constituents of abused "bath salts" preparations and represent a large family of structurally related compounds that function as cocaine-like inhibitors or amphetamine-like substrates of dopamine (DAT), norepinephrine (NET), and serotonin (SERT) transporters. Preclinical evidence suggests that some cathinones (e.g., MDPV and α-PVP) are more effective reinforcers than prototypical stimulant drugs of abuse, such as cocaine or methamphetamine. Although the reinforcing potency of these cathinones is related to their potency to inhibit DAT, less is known about the pharmacological determinants of their unusually high reinforcing effectiveness. To this end, we tested the hypothesis that reinforcing effectiveness of cathinone stimulants is positively correlated with their selectivity for DAT relative to SERT. Uptake inhibition assays in rat brain synaptosomes were used to directly compare the potency of MDPV, MDPBP, MDPPP, α-PVP, α-PPP, and cocaine at DAT, NET, and SERT, whereas intravenous self-administration in rats was used to quantify relative reinforcing effectiveness of the drugs using progressive ratio (PR) and behavioral economic procedures. All cathinones were more potent at DAT than NET or SERT, with a rank order for selectivity at DAT over SERT of α-PVP > α-PPP > MDPV > MDPBP > MDPPP > cocaine. These synthetic cathinones were more effective reinforcers than cocaine, and the measures of reinforcing effectiveness determined by PR and demand curve analyses were highly correlated with selectivity for DAT over SERT. Together, these studies provide strong and convergent evidence that the abuse potential of stimulant drugs is mediated by uptake inhibition at DAT, with activity at SERT serving as a negative modulator of reinforcing effectiveness.

Reactive oxygen species (ROS) and compromised antioxidant defense may contribute to brain disorders such as stroke, amyotrophic lateral sclerosis, etc. Nitroxides are redox-sensitive paramagnetic contrast agents and antioxidants. The ability of a blood-brain barrier (BBB)-permeable nitroxide, methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (MC-P), as a magnetic resonance-imaging (MRI) contrast agent for brain tissue redox imaging was tested. MC-P relaxation in rodent brain was quantified by MRI using a fast Look-Locker T 1 -mapping sequence. In the cerebral cortex and thalamus, the MRI signal intensity increased up to 50% after MC-P injection, but increased only by 2.7% when a BBB-impermeable nitroxide, 3CxP (3-carboxy-2,2,5,5,5-tetramethylpyrrolidine-1-oxyl) was used. The maximum concentrations in the thalamus and cerebral cortex after MC-P injection were calculated to be 1.9±0.35 and 3.0±0.50 mmol/L, respectively. These values were consistent with the ex vivo data of brain tissue and blood concentration obtained by electron paramagnetic resonance (EPR) spectroscopy. Also, reduction rates of MC-P were significantly decreased after reperfusion following transient MCAO (middle cerebral artery occlusion), a condition associated with changes in redox status resulting from oxidative damage. These results show the use of BBB-permeable nitroxides as MRI contrast agents and antioxidants to evaluate the role of ROS in neurologic diseases.

"Bath salts" preparations contain synthetic cathinones which interact with monoamine transporters and function as either monoamine uptake inhibitors or releasers. 3,4-Methylenedioxypyrovalerone (MDPV), 3,4-methylenedioxymethcathinone (methylone), and 4-methylmethcathinone (mephedrone) were three of the most common cathinones (i.e., "first-generation" cathinones); however, after the US Drug Enforcement Administration placed them under Schedule I regulations, they were replaced with structurally related cathinones that were not subject to regulations (i.e., "second-generation" cathinones). Although the reinforcing effects of some second-generation cathinones have been described (e.g., α-pyrrolidinopentiophenone [α-PVP]), little is known about how structural modifications, particularly those involving the methylenedioxy moiety and α-alkyl side chain, impact the abuse liability of other second-generation cathinones (e.g., α-pyrrolidinopropiophenone [α-PPP], 3,4-methylenedioxy-α-pyrrolidinobutiophenone [MDPBP], and 3,4-methylenedioxy-α-pyrrolidinopropiophenone [MDPPP]). The present study used male Sprague-Dawley rats (n = 12 per drug) to directly compare: (1) the acquisition of responding for α-PVP (0.032 mg/kg/inf), α-PPP (0.32 mg/kg/inf), MDPBP (0.1 mg/kg/inf), and MDPPP (0.32 mg/kg/inf) under a fixed ratio (FR) 1 schedule of reinforcement; and (2) full dose-response curves for each drug to maintain responding under an FR5 schedule of reinforcement. The average number of days (∼4 days) and percentage (100%) of rats that acquired self-administration was similar for each drug. The observed rank order potency to maintain responding under an FR5 schedule of reinforcement (α-PVP ≈ MDPBP>α-PPP > MDPPP) is consistent with their potencies to inhibit dopamine uptake. These are the first studies to report on the reinforcing effects of the unregulated second-generation cathinones MDPBP, MDPPP, and α-PPP and indicate all three compounds are readily self-administered, suggesting each possesses high potential for abuse. This article is part of the Special Issue entitled 'Designer Drugs and Legal Highs.'

2,5-Dimethoxyphenethylamines (2C compounds) are 5-HT receptor agonists that induce hallucinogenic effects. N-methoxybenzylation of 2C compounds markedly increases their affinity for 5-HT receptors, and two such analogs, 2-(4-chloro-2,5-dimethoxyphenyl)-N-[(2-methoxyphenyl)methyl]ethanamine (25C-NBOMe) and 2-(4-iodo-2,5-dimethoxyphenyl)-N-[(2-methoxyphenyl)methyl]ethanamine (25I-NBOMe), have emerged in recreational drug markets. Here, we investigated the neuropharmacology of 25C-NBOMe and 25I-NBOMe in rats, as compared to their 2C analogs and the prototypical 5-HT agonist 1-(4-iodo-2,5-dimethoxyphenyl)propan-2-amine (DOI). Compounds were tested in vitro using 5-HT receptor binding and calcium mobilization assays. For in vivo experiments, 25C-NBOMe (0.01-0.3 mg/kg), 25I-NBOMe (0.01-0.3 mg/kg), 2-(4-chloro-2,5-dimethoxyphenyl)ethanamine (2C-C) (0.1-3.0 mg/kg), 2-(4-iodo-2,5-dimethoxyphenyl)ethanamine (2C-I) (0.1-3.0 mg/kg) and DOI (0.03-1.0 mg/kg) were administered subcutaneously (sc) to male rats, and 5-HT-mediated behaviors were assessed. NBOMes displayed higher affinity for 5-HT receptors than their 2C counterparts but were substantially weaker in functional assays. 25C-NBOMe and 25I-NBOMe were much more potent at inducing wet dog shakes (WDS) and back muscle contractions (BMC) when compared to 2C-C and 2C-I. Pretreatment with the selective 5-HT antagonist (R)-(2,3-dimethoxyphenyl){1-[2-(4-fluorophenyl)ethyl]-4-piperidinyl}methanol (M100907) reversed behaviors produced by all agonists. Interestingly, binding affinities at the 5-HT receptor were significantly correlated with potencies to induce BMC but not WDS. Our findings show that NBOMes are highly potent 5-HT agonists in rats, similar to effects in mice, and consistent with the reported hallucinogenic effects in human users. This article is part of the Special Issue entitled 'Psychedelics: New Doors, Altered Perceptions'.

Active immunization is an emerging potential modality to combat fatal overdose amid the opioid epidemic. In this study, we described the design, synthesis, formulation, and animal testing of an efficacious vaccine against fentanyl. The vaccine formulation is composed of a novel fentanyl hapten conjugated to tetanus toxoid (TT) and adjuvanted with liposomes containing monophosphoryl lipid A adsorbed on aluminum hydroxide. The linker and hapten N -phenyl- N -(1-(4-(3-(tritylthio)propanamido)phenethyl)piperidin-4-yl)propionamide were conjugated sequentially to TT using amine- N -hydroxysuccinimide-ester and thiol–maleimide reaction chemistries, respectively. Conjugation was facile, efficient, and reproducible with a protein recovery of >98% and a hapten density of 30–35 per carrier protein molecule. In mice, immunization induced high and robust antibody endpoint titers in the order of >10 6 against the hapten. The antisera bound fentanyl, carfentanil, cyclopropyl fentanyl, para -fluorofentanyl, and furanyl fentanyl in vitro with antibody-drug dissociation constants in the range of 0.36–4.66 nM. No cross-reactivity to naloxone, naltrexone, methadone, or buprenorphine was observed. In vivo , immunization shifted the antinociceptive dose–response curve of fentanyl to higher doses. Collectively, these preclinical results showcased the desired traits of a potential vaccine against fentanyl and demonstrated the feasibility of immunization to combat fentanyl-induced effects.

Low‐efficacy mu‐opioid receptor (MOR) agonists represent promising therapeutics, but existing compounds (e.g., buprenorphine, nalbuphine) span a limited range of low MOR efficacies and have poor MOR selectivity. Accordingly, new and selective low‐efficacy MOR agonists are of interest. A novel set of chiral C9‐substituted phenylmorphans has been reported to display improved MOR selectivity and a range of high‐to‐low MOR efficacies under other conditions; however, a full opioid receptor binding profile for these drugs has not been described. Additionally, studies in mice will be useful for preclinical characterization of these novel compounds, but the pharmacology of these drugs in mice has also not been examined. Accordingly, the present study characterized the binding selectivity and in vitro efficacy of these compounds using assays of opioid receptor binding and ligand‐stimulated [35S]GTPɣS binding. Additionally, locomotor effects were evaluated as a first step for in vivo behavioral assessment in mice. The high‐efficacy MOR agonist and clinically effective antidepressant tianeptine was included as a comparator. In binding studies, all phenylmorphans showed improved MOR selectivity relative to existing lower‐efficacy MOR agonists. In the ligand‐stimulated [35S]GTPɣS binding assay, seven phenylmorphans had graded levels of sub‐buprenorphine MOR efficacy. In locomotor studies, the compounds again showed graded efficacy with a rapid onset and ≥1 h duration of effects, evidence for MOR mediation, and minor sex differences. Tianeptine functioned as a high‐efficacy MOR agonist. Overall, these in vitro and in vivo studies support the characterization of these compounds as MOR‐selective ligands with graded MOR efficacy and utility for further behavioral studies in mice.

Receptor-type protein tyrosine phosphatase D (PTPRD) is a neuronal cell-adhesion molecule/synaptic specifier that has been implicated in addiction vulnerability and stimulant reward by human genomewide association and mouse cocaine-conditioned place-preference data. However, there have been no reports of effects of reduced expression on cocaine self-administration. There have been no reports of PTPRD targeting by any small molecule. There are no data about behavioral effects of any PTPRD ligand. We now report (i) robust effects of heterozygous PTPRD KO on cocaine self-administration (These data substantially extend prior conditioned place-preference data and add to the rationale for PTPRD as a target for addiction therapeutics.); (ii) identification of 7-butoxy illudalic acid analog (7-BIA) as a small molecule that targets PTPRD and inhibits its phosphatase with some specificity; (iii) lack of toxicity when 7-BIA is administered to mice acutely or with repeated dosing; (iv) reduced cocaine-conditioned place preference when 7-BIA is administered before conditioning sessions; and (v) reductions in well-established cocaine self-administration when 7-BIA is administered before a session (in WT, not PTPRD heterozygous KOs). These results add to support for PTPRD as a target for medications to combat cocaine use disorders. 7-BIA provides a lead compound for addiction therapeutics.