RESUMO
The kappa opioid receptor has been identified as a promising therapeutic target for promoting remyelination. In the current study, we evaluated the ability of nalfurafine to promote oligodendrocyte progenitor cell (OPC) differentiation and myelination in vitro, and its efficacy in an extended, cuprizone-induced demyelination model. Primary mouse (C57BL/6J) OPC-containing cultures were treated with nalfurafine (0.6-200 nM), clemastine (0.01-100 µM), T3 (30 ng/mL), or vehicle for 5 days. Using immunocytochemistry and confocal microscopy, we found that nalfurafine treatment increased OPC differentiation, oligodendrocyte (OL) morphological complexity, and myelination of nanofibers in vitro. Adult male mice (C57BL/6J) were given a diet containing 0.2% cuprizone and administered rapamycin (10 mg/kg) once daily for 12 weeks followed by 6 weeks of treatment with nalfurafine (0.01 or 0.1 mg/kg), clemastine (10 mg/kg), or vehicle. We quantified the number of OLs using immunofluorescence, gross myelination using black gold staining, and myelin thickness using electron microscopy. Cuprizone + rapamycin treatment produced extensive demyelination and was accompanied by a loss of mature OLs, which was partially reversed by therapeutic administration of nalfurafine. We also assessed these mice for functional behavioral changes in open-field, horizontal bar, and mouse motor skill sequence tests (complex wheel running). Cuprizone + rapamycin treatment resulted in hyperlocomotion, poorer horizontal bar scores, and less distance traveled on the running wheels. Partial recovery was observed on both the horizontal bar and complex running wheel tests over time, which was facilitated by nalfurafine treatment. Taken together, these data highlight the potential of nalfurafine as a remyelination-promoting therapeutic.
Assuntos
Cuprizona , Doenças Desmielinizantes , Camundongos Endogâmicos C57BL , Morfinanos , Bainha de Mielina , Sirolimo , Compostos de Espiro , Animais , Morfinanos/farmacologia , Masculino , Compostos de Espiro/farmacologia , Doenças Desmielinizantes/induzido quimicamente , Doenças Desmielinizantes/patologia , Doenças Desmielinizantes/tratamento farmacológico , Camundongos , Bainha de Mielina/efeitos dos fármacos , Bainha de Mielina/patologia , Bainha de Mielina/metabolismo , Sirolimo/farmacologia , Cuprizona/toxicidade , Células Cultivadas , Modelos Animais de Doenças , Células Precursoras de Oligodendrócitos/efeitos dos fármacos , Células Precursoras de Oligodendrócitos/metabolismo , Diferenciação Celular/efeitos dos fármacosRESUMO
The synthesis of stereochemically pure oximes, amines, saturated and unsaturated cyanomethyl compounds, and methylaminomethyl compounds at the C9 position in 3-hydroxy-N-phenethyl-5-phenylmorphans provided µ-opioid receptor (MOR) agonists with varied efficacy and potency. One of the most interesting compounds, (2-((1S,5R,9R)-5-(3-hydroxyphenyl)-2-phenethyl-2-azabicyclo[3.3.1]nonan-9-yl)acetonitrile), was found to be a potent partial MOR agonist (EC50 = 2.5 nM, %Emax = 89.6%), as determined in the forskolin-induced cAMP accumulation assay. Others ranged in potency and efficacy at the MOR, from nanomolar potency with a C9 cyanomethyl compound (EC50 = 0.85 nM) to its totally inactive diastereomer, and three compounds exhibited weak MOR antagonist activity (the primary amine 3, the secondary amine 8, and the cyanomethyl compound 41). Many of the compounds were fully efficacious; their efficacy and potency were affected by both the stereochemistry of the molecule and the specific C9 substituent. Most of the MOR agonists were selective in their receptor interactions, and only a few had δ-opioid receptor (DOR) or κ-opioid receptor (KOR) agonist activity. Only one compound, a C9-methylaminomethyl-substituted phenylmorphan, was moderately potent and fully efficacious as a KOR agonist (KOR EC50 = 18 nM (% Emax = 103%)).
Assuntos
Aminas , Oximas , Oximas/química , Oximas/farmacologia , Estereoisomerismo , Relação Estrutura-Atividade , Aminas/química , Aminas/farmacologia , Receptores Opioides mu/metabolismo , Receptores Opioides mu/agonistas , Humanos , Animais , Estrutura Molecular , Células CHO , Morfinanos/química , Morfinanos/farmacologiaRESUMO
The 5-(3-hydroxy)phenylmorphan structural class of compounds are unlike the classical morphinans, 4,5-epoxymorphinans, and 6,7-benzomorphans, in that they have an equatorially oriented aromatic ring rather than the axial orientation of that ring found in the classical opioids. This modified and simplified opioid-like structure has been shown to retain antinociceptive activity, depending on its stereochemistry and substituents, and some of them have been found to be much more potent than morphine. A simple C9-hydroxy-5-(3-hydroxy)phenylmorphan enantiomer was found to be about 500 times more potent than morphine in vivo. We have previously examined C9-alkenyl and hydroxyalkyl substituents in the N-phenethyl-5-(3-hydroxy)phenylmorphan class of compounds. Comparable C9-alkyl (methyl through butyl) substituents, with their sets of diastereomers, have not been explored. All these compounds have now been synthesized to determine the effect chain-length and stereochemistry at the C9 position in the molecule might have on their interaction with opioid receptors. We now report the synthesis and in vitro activity of 16 compounds, the C9-methyl, ethyl, propyl, and butyl diastereomers, using the inhibition of forskolin-induced cAMP accumulation assay. Several potent (sub-nanomolar and nanomolar) MOR compounds were found to be selective agonists with varying efficacy. Of greatest interest, a selective MOR antagonist was discovered; it did not display any DOR or KOR agonist activity in vitro, was three times more potent than naltrexone, and was found to antagonize the EC90 of fentanyl at MOR to a greater extent than naltrexone.
Assuntos
Morfinanos , Receptores Opioides mu , Receptores Opioides mu/química , Naltrexona/farmacologia , Relação Estrutura-Atividade , Morfinanos/química , Morfina , Analgésicos Opioides/farmacologiaRESUMO
Kappa opioid receptor (KOR) agonists have preclinical antipsychostimulant effects; however, adverse side effects have limited their therapeutic development. In this preclinical study, conducted in Sprague Dawley rats, B6-SJL mice, and non-human primates (NHPs), we evaluated the G-protein-biased analogue of salvinorin A (SalA), 16-bromo salvinorin A (16-BrSalA), for its anticocaine effects, side effects, and activation of cellular signaling pathways. 16-BrSalA dose-dependently decreased the cocaine-primed reinstatement of drug-seeking behavior in a KOR-dependent manner. It also decreased cocaine-induced hyperactivity, but had no effect on responding for cocaine on a progressive ratio schedule. Compared to SalA, 16-BrSalA had an improved side effect profile, with no significant effects in the elevated plus maze, light-dark test, forced swim test, sucrose self-administration, or novel object recognition; however, it did exhibit conditioned aversive effects. 16-BrSalA increased dopamine transporter (DAT) activity in HEK-293 cells coexpressing DAT and KOR, as well as in rat nucleus accumbens and dorsal striatal tissue. 16-BrSalA also increased the early phase activation of extracellular-signal-regulated kinases 1 and 2, as well as p38 in a KOR-dependent manner. In NHPs, 16-BrSalA caused dose-dependent increases in the neuroendocrine biomarker prolactin, similar to other KOR agonists, at doses without robust sedative effects. These findings highlight that G-protein-biased structural analogues of SalA can have improved pharmacokinetic profiles and fewer side effects while maintaining their anticocaine effects.
Assuntos
Cocaína , Camundongos , Ratos , Humanos , Animais , Cocaína/farmacologia , Receptores Opioides kappa/metabolismo , Ratos Sprague-Dawley , Células HEK293 , Ansiedade/tratamento farmacológico , Recompensa , LocomoçãoRESUMO
All possible diastereomeric C9-hydroxymethyl-, hydroxyethyl-, and hydroxypropyl-substituted 5-phenylmorphans were synthesized to explore the three-dimensional space around the C9 substituent in our search for potent MOR partial agonists. These compounds were designed to lessen the lipophilicity observed with their C9-alkenyl substituted relatives. Many of the 12 diastereomers that were obtained were found to have nanomolar or subnanomolar potency in the forskolin-induced cAMP accumulation assay. Almost all these potent compounds were fully efficacious, and three of those chosen for in vivo evaluation, 15, 21, and 36, were all extremely G-protein biased; none of the three compounds recruited beta-arrestin2. Only one of the 12 diastereomers, 21 (3-((1S,5R,9R)-9-(2-hydroxyethyl)-2-phenethyl-2-azabicyclo[3.3.1]nonan-5-yl)phenol), was a MOR partial agonist with good, but not full, efficacy (Emax = 85%) and subnanomolar potency (EC50 = 0.91 nM) in the cAMP assay. It did not have any KOR agonist activity. This compound was unlike morphine in that it had a limited ventilatory effect in vivo. The activity of 21 could be related to one or more of three well-known theories that attempt to predict a dissociation of the desired analgesia from the undesirable opioid-like side-effects associated with clinically used opioids. In accordance with the theories, 21 was a potent MOR partial agonist, it was highly G-protein biased and did not attract beta-arrestin2, and it was found to have both MOR and DOR agonist activity. All the other diastereomers that were synthesized were either much less potent than 21 or had either too little or too much efficacy for our purposes. It was also noted that a C9-methoxymethyl compound with 1R,5S,9R stereochemistry (41) was more potent than the comparable C9-hydroxymethyl compound 11 (EC50 = 0.65 nM for 41 vs. 2.05 nM for 11). Both 41 and 11 were fully efficacious.
Assuntos
Morfinanos , Receptores Opioides mu , Morfinanos/química , Morfina , Analgésicos Opioides/químicaRESUMO
Pseudomonas aeruginosa is an increasingly antibiotic-resistant pathogen that causes severe lung infections, burn wound infections, and diabetic foot infections. P. aeruginosa produces the siderophore pyochelin through the use of a non-ribosomal peptide synthetase (NRPS) biosynthetic pathway. Targeting members of siderophore NRPS proteins is one avenue currently under investigation for the development of new antibiotics against antibiotic-resistant organisms. Here, the crystal structure of the pyochelin adenylation domain PchD is reported. The structure was solved to 2.11 Å when co-crystallized with the adenylation inhibitor 5'-O-(N-salicylsulfamoyl)adenosine (salicyl-AMS) and to 1.69 Å with a modified version of salicyl-AMS designed to target an active site cysteine (4-cyano-salicyl-AMS). In the structures, PchD adopts the adenylation conformation, similar to that reported for AB3403 from Acinetobacter baumannii.
Assuntos
Pseudomonas aeruginosa , Sideróforos , Antibacterianos/metabolismo , Antibacterianos/farmacologia , Fenóis , Pseudomonas aeruginosa/metabolismo , Salicilatos/metabolismo , Sideróforos/química , TiazóisRESUMO
The design of enantiopure stereoisomers of N-2-phenylcyclopropylmethyl-substituted ortho-c oxide-bridged phenylmorphans, the E and Z isomers of an N-cinnamyl moiety, and N-propyl enantiomers were based on combining the most potent oxide-bridged phenylmorphan (the ortho-c isomer) with the most potent N-substituent that we previously found with a 5-(3-hydroxy)phenylmorphan (i.e., N-2-phenylcyclopropyl methyl moieties, N-cinnamyl, and N-propyl substituents). The synthesis of the eight enantiopure N-2-phenylcyclopropylmethyl ortho-c oxide-bridged phenylmorphans and six additional enantiomers of the N-substituted ortho-c oxide-bridged phenylmorphans (N-E and Z-cinnamyl compounds, and N-propyl compounds) was accomplished. The synthesis started from common intermediates (3R,6aS,11aS)-10-methoxy-1,3,4,5,6,11a-hexahydro-2H-3,6a-methano-benzofuro[2,3-c]azocine (+)-6 and its enantiomer, (3S, 6aR, 11aR)-(-)-6, respectively. The enantiomers of ±-6 were obtained through salt formation with (S)-(+)- and (R)-(-)-p-methylmandelic acid, and the absolute configuration of the (R)-(-)-p-methylmandelate salt of (3S, 6aR, 11aR)-(-)-6 was determined by single-crystal X-ray analysis. The enantiomeric secondary amines were reacted with N-(2-phenylcyclopropyl)methyl derivatives, 2-(E)-cinnamyl bromide, and (Z)-3-phenylacrylic acid. These products led to all of the desired N-derivatives of the ortho-c oxide-bridged phenylmorphans. Their opioid receptor binding affinity was measured. The compounds with MOR affinity < 50 nM were examined for their functional activity in the forskolin-induced cAMP accumulation assay. Only the enantiomer of the N-phenethyl ortho-c oxide-bridged phenylmorphan ((-)-1), and only the (3S,6aR,11aR)-2-(((1S,2S)-2-phenylcyclopropyl)methyl)-1,3,4,5,6,11a-hexahydro-2H-3,6a-methanobenzofuro[2,3-c]azocin-10-ol isomer ((+)-17), and the N-phenylpropyl derivative ((-)-25) had opioid binding affinity < 50 nM. Both (-)-1 and (-)-25 were partial agonists in the cAMP assay, with the former showing high potency and low efficacy, and the latter with lower potency and less efficacy. Most interesting was the N-2-phenylcyclopropylmethyl (3S,6aR,11aR)-2-(1S,2S)-enantiomer ((+)-17). That compound had good MOR binding affinity (Ki = 11.9 nM) and was found to have naltrexone-like potency as a MOR antagonist (IC50 = 6.92 nM).
Assuntos
Morfinanos , Óxidos , Cristalografia por Raios X , Óxidos/química , Morfinanos/química , Isomerismo , Receptores Opioides muRESUMO
Four sets of diastereomeric C9-alkenyl 5-phenylmorphans, varying in the length of the C9-alkenyl chain, were designed to examine the effect of these spatially distinct ligands on opioid receptors. Functional activity was obtained by forskolin-induced cAMP accumulation assays and several compounds were examined in the [35S]GTPgS assay and in an assay for respiratory depression. In each of the four sets, similarities and differences were observed dependent on the length of their C9-alkenyl chain and, most importantly, their stereochemistry. Three MOR antagonists were found to be as or more potent than naltrexone and, unlike naltrexone, none had MOR, KOR, or DOR agonist activity. Several potent MOR full agonists were obtained, and, of particular interest partial agonists were found that exhibited less respiratory depression than that caused by morphine. The effect of stereochemistry and the length of the C9-alkenyl chain was also explored using molecular modeling. The MOR antagonists were found to interact with the inactive (4DKL) MOR crystal structures and agonists were found to interact with the active (6DDF) MOR crystal structures. The comparison of their binding modes at the mouse MOR was used to gain insight into the structural basis for their stereochemically induced pharmacological differences.
Assuntos
Naltrexona , Insuficiência Respiratória , Animais , Células CHO , Colforsina , Cricetinae , Ligantes , Camundongos , Morfina/farmacologia , Receptores Opioides/metabolismo , Receptores Opioides delta/metabolismo , Receptores Opioides mu/metabolismoRESUMO
BACKGROUND: Synthetic cathinones display overlapping behavioral effects with psychostimulants (e.g., methamphetamine [MA]) and/or entactogens (e.g., 3,4-methylenedioxymethaphetamine [MDMA])-presumably reflecting their dopaminergic and/or serotonergic activity. The discriminative stimulus effects of MDMA thought to be mediated by such activity have been well characterized in rodents but have not been fully examined in nonhuman primates. METHODS: The present studies were conducted to systematically evaluate the discriminative stimulus effects of 5 abused synthetic cathinones (methylenedioxypyrovalerone [MDPV], α-pyrrolidinovalerophenone [α-PVP], methcathinone [MCAT], mephedrone, and methylone) in adult male squirrel monkeys trained to distinguish intramuscular injections of MA (0.1 mg/kg; n = 4) or MDMA (0.6 mg/kg; n = 4) from vehicle. RESULTS: Each training drug produced dose-dependent effects and, at the highest dose, full substitution. MDMA produced predominantly vehicle-like responding in the MA-trained group, whereas the highest dose of MA (0.56 mg/kg) produced partial substitution (approximately 90% appropriate lever responding in one-half of the subjects) in the MDMA-trained group. MDPV, α-PVP, and MCAT produced full substitution in MA-trained subjects, but, at the same or higher doses, only substituted for MDMA in one-half of the subjects, consistent with primarily dopaminergically mediated interoceptive effects. In contrast, mephedrone and methylone fully substituted in MDMA-trained subjects but failed to fully substitute for the training drug in MA-trained subjects, suggesting a primary role for serotonergic actions in their interoceptive effects. CONCLUSIONS: These findings suggest that differences in the interoceptive effects of synthetic cathinones in nonhuman primates reflect differing compositions of monoaminergic actions that also may mediate their subjective effects in humans.
Assuntos
Alcaloides/farmacologia , Estimulantes do Sistema Nervoso Central/farmacologia , Aprendizagem por Discriminação/efeitos dos fármacos , Interocepção/efeitos dos fármacos , Metanfetamina/farmacologia , N-Metil-3,4-Metilenodioxianfetamina/farmacologia , Psicotrópicos/farmacologia , Alcaloides/administração & dosagem , Animais , Comportamento Animal/efeitos dos fármacos , Benzodioxóis/farmacologia , Estimulantes do Sistema Nervoso Central/administração & dosagem , Masculino , Metanfetamina/administração & dosagem , Metanfetamina/análogos & derivados , N-Metil-3,4-Metilenodioxianfetamina/administração & dosagem , Propiofenonas/farmacologia , Psicotrópicos/administração & dosagem , Pirrolidinas/farmacologia , Saimiri , Catinona SintéticaRESUMO
BACKGROUND: One of the most prominent opioid analgesics in the United States is the high potency agonist fentanyl. It is used in the treatment of acute and chronic pain and as an anesthetic adjuvant. When used inappropriately, however, ingestion of just a few milligrams of fentanyl or other synthetic opioid can cause opioid-induced respiratory depression (OIRD), often leading to death. Currently, the treatment of choice for OIRD is the opioid receptor antagonist naloxone. Recent reports, however, suggest that higher doses or repeated dosing of naloxone (due to recurrence of respiratory depression) may be required to reverse fully fentanyl-induced respiratory depression, rendering this treatment inadequate. To combat this synthetic opioid overdose crisis, this research aims at identifying a novel opioid reversal agent with enhanced efficacy towards fentanyl and other synthetic opioids. METHODS: A series of naltrexone analogues were characterized for their ability to antagonize the effects of fentanyl in vitro utilizing a modified forskolin-induced cAMP accumulation assay. Lead analogue 29 was chosen to undergo further PK studies, followed by in vivo pharmacological analysis to determine its ability to antagonize opioid-induced antinociception in the hot plate assay. RESULTS: A series of potent MOR antagonists were identified, including the highly potent analogue 29 (IC50 = 2.06 nM). Follow-up PK studies revealed 29 to possess near 100% bioavailability following IP administration. Brain concentrations of 29 surpassed plasma concentrations, with an apparent terminal half-life of ~ 80 min in mice. In the hot plate assay, 29 dose-dependently (0.01-0.1 mg/kg; IP) and fully antagonized the antinociception induced by oxycodone (5.6 mg/kg; IP). Furthermore, the dose of 29 that is fully effective in preventing oxycodone-induced antinociception (0.1 mg/kg) was ineffective against locomotor deficits caused by the KOR agonist U50,488. CONCLUSIONS: Methods have been developed that have utility to identify enhanced rescue agents for the treatment of OIRD. Analogue 29, possessing potent MOR antagonist activity in vitro and in vivo, provides a promising lead in our search for an enhanced synthetic opioid rescue agent.
Assuntos
Analgésicos Opioides/efeitos adversos , Fentanila/efeitos adversos , Naltrexona , Antagonistas de Entorpecentes , Animais , Desenho de Fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Naltrexona/síntese química , Naltrexona/farmacocinética , Naltrexona/farmacologia , Antagonistas de Entorpecentes/síntese química , Antagonistas de Entorpecentes/farmacocinética , Antagonistas de Entorpecentes/farmacologiaRESUMO
Herpes simplex virus 1 (HSV-1) has infected more than 80% of the population. Reactivation of the virus causes diseases ranging in severity from benign cold sores to fatal encephalitis. Current treatments involve viral DNA replication inhibitors, but the emergence of drug-resistant mutants is observed frequently, highlighting the need for novel antiviral therapies. Infected cell protein 0 (ICP0) of HSV-1 is encoded by an immediate early gene and plays a fundamental role during infection, because it enables viral gene expression and blocks antiviral responses. One mechanism by which ICP0 functions is through an E3 ubiquitin ligase activity that induces the degradation of targeted proteins. A ΔICP0 virus or mutants with deficiencies in E3 ligase activity cannot counteract beta interferon (IFN-ß)-induced restriction of viral infection, are highly immunogenic, are avirulent, and fail to spread. Thus, small molecules interfering with essential and conserved ICP0 functions are expected to compromise HSV-1 infection. We have developed a high-throughput screening assay, based on the autoubiquitination properties of ICP0, to identify small-molecule inhibitors of ICP0 E3 ubiquitin ligase activity. Through a pilot screening procedure, we identified nine compounds that displayed dose-dependent inhibitory effects on ICP0 but not on Mdm2, a control E3 ubiquitin ligase. Following validation, one compound displayed ICP0-dependent inhibition of HSV-1 infection. This compound appeared to bind ICP0 in a cellular thermal shift assay, it blocked ICP0 self-elimination, and it blocked wild-type but not ICP0-null virus gene expression. This scaffold displays specificity and could be used to develop optimized ICP0 E3 ligase inhibitors.IMPORTANCE Since acyclovir and its derivatives were launched for herpesviruses control almost four decades ago, the search for novel antivirals has waned. However, as human life expectancy has increased, so has the number of immunocompromised individuals who receive prolonged treatment for HSV recurrences. This has led to an increase in unresponsive patients due to acquired viral drug resistance. Thus, novel treatments need to be explored. Here we explored the HSV-1 ICP0 E3 ligase as a potential antiviral target because (i) ICP0 is expressed before virus replication, (ii) it is essential for infection in vivo, (iii) it is required for efficient reactivation of the virus from latency, (iv) inhibition of its E3 ligase activity would sustain host immune responses, and (v) it is shared by other herpesviruses. We report a compound that inhibits HSV-1 infection in an ICP0-dependent manner by inhibiting ICP0 E3 ligase activity.
Assuntos
Herpesvirus Humano 1/efeitos dos fármacos , Herpesvirus Humano 1/metabolismo , Ensaios de Triagem em Larga Escala , Proteínas Imediatamente Precoces/efeitos dos fármacos , Proteínas Imediatamente Precoces/metabolismo , Ubiquitina-Proteína Ligases/efeitos dos fármacos , Linhagem Celular , Replicação do DNA , Regulação Viral da Expressão Gênica , Herpesvirus Humano 1/genética , Interações Hospedeiro-Patógeno , Humanos , Proteínas Imediatamente Precoces/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Proteínas Virais , Replicação Viral/efeitos dos fármacosRESUMO
Mu-opioid receptor (MOR) agonists are highly efficacious for the treatment of pain but have significant abuse liability. Recently, we reported that nalfurafine, when combined with oxycodone at a certain ratio, reduced the reinforcing effects of oxycodone in rats while producing additive antinociceptive effects. Questions remain, however, including if the combination will function as a reinforcer in drug-naïve rats, and if the combination produces aversive effects that could explain nalfurafine's ability to reduce oxycodone self-administration? In the present study, we investigated nalfurafine's ability to reduce acquisition of oxycodone self-administration when the two were self-administered as a mixture in drug-naïve rats and nalfurafine's ability to attenuate a conditioned place preference (CPP) induced by oxycodone. In the self-administration study, male Sprague-Dawley rats self-administered intravenous injections of oxycodone (0.056 mg/kg/injection), an oxycodone/nalfurafine combination (0.056/0.0032 mg/kg/injection), or saline under fixed-ratio schedules of reinforcement for 20 days to compare rates of acquisition of drug taking. In the CPP assay, male Sprague-Dawley rats received subcutaneous injections of either saline, oxycodone (3.2 mg/kg), nalfurafine (0.18 mg/kg), or an oxycodone/nalfurafine combination at the same ratio used in the self-administration study (3.2 mg/kg/0.18 mg/kg). All subjects self-administering oxycodone alone met acquisition criteria. However, only 13% of subjects self-administering oxycodone/nalfurafine met criteria, and no subjects acquired self-administration of saline. Oxycodone, but not nalfurafine alone or the oxycodone/nalfurafine combination, produced rewarding effects in rats in the CPP test. These findings suggest that the combination of oxycodone and nalfurafine will be less habit forming in opioid-naïve patients than oxycodone alone.
Assuntos
Morfinanos/farmacologia , Transtornos Relacionados ao Uso de Opioides/tratamento farmacológico , Receptores Opioides kappa/agonistas , Compostos de Espiro/farmacologia , Analgésicos Opioides/farmacologia , Animais , Condicionamento Clássico/efeitos dos fármacos , Relação Dose-Resposta a Droga , Masculino , Morfinanos/metabolismo , Transtornos Relacionados ao Uso de Opioides/prevenção & controle , Oxicodona/efeitos adversos , Oxicodona/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores Opioides/agonistas , Receptores Opioides/metabolismo , Receptores Opioides kappa/metabolismo , Reforço Psicológico , Recompensa , Autoadministração , Compostos de Espiro/metabolismo , Transtornos Relacionados ao Uso de Substâncias/tratamento farmacológico , Transtornos Relacionados ao Uso de Substâncias/prevenção & controleRESUMO
Small molecule agonists of TLR7/8, such as imidazoquinolines, are validated agonists for the treatment of cancer and for use in vaccine adjuvants. Imidazoquinolines have been extensively modified to understand the structure-activity relationship (SAR) at the N1- and C2-positions resulting in the clinical drug imiquimod, resiquimod, and several other highly potent analogues. However, the SAR of the aryl ring has not been fully elucidated in the literature. This initial study examines the SAR of C7-substituted imidazoquinolines. These compounds not only demonstrated that TLR7/8 tolerate changes at the C7-position but can increase potency and change their cytokine profiles. The most notable TLR7/8 agonists developed from this study 5, 8, and 14 which are up to 4-fold and 2-fold more active than resiquimod for TLR8 and/or TLR7, respectively, and up to 100-fold more active than the FDA approved imiquimod for TLR7.
Assuntos
Quinolinas/química , Receptor 7 Toll-Like/agonistas , Receptor 8 Toll-Like/agonistas , Sítios de Ligação , Citocinas/metabolismo , Humanos , Imidazóis/química , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Simulação de Acoplamento Molecular , Quinolinas/metabolismo , Relação Estrutura-Atividade , Receptor 7 Toll-Like/metabolismo , Receptor 8 Toll-Like/metabolismoRESUMO
(-)-N-Phenethyl analogs of optically pure N-norhydromorphone were synthesized and pharmacologically evaluated in several in vitro assays (opioid receptor binding, stimulation of [35S]GTPγS binding, forskolin-induced cAMP accumulation assay, and MOR-mediated ß-arrestin recruitment assays). "Body" and "tail" interactions with opioid receptors (a subset of Portoghese's message-address theory) were used for molecular modeling and simulations, where the "address" can be considered the "body" of the hydromorphone molecule and the "message" delivered by the substituent (tail) on the aromatic ring of the N-phenethyl moiety. One compound, N-p-chloro-phenethynorhydromorphone ((7aR,12bS)-3-(4-chlorophenethyl)-9-hydroxy-2,3,4,4a,5,6-hexahydro-1H-4,12-methanobenzofuro[3,2-e]isoquinolin-7(7aH)-one, 2i), was found to have nanomolar binding affinity at MOR and DOR. It was a potent partial agonist at MOR and a full potent agonist at DOR with a δ/µ potency ratio of 1.2 in the ([35S]GTPγS) assay. Bifunctional opioids that interact with MOR and DOR, the latter as agonists or antagonists, have been reported to have fewer side-effects than MOR agonists. The p-chlorophenethyl compound 2i was evaluated for its effect on respiration in both mice and squirrel monkeys. Compound 2i did not depress respiration (using normal air) in mice or squirrel monkeys. However, under conditions of hypercapnia (using air mixed with 5% CO2), respiration was depressed in squirrel monkeys.
Assuntos
Hidromorfona/análogos & derivados , Hipercapnia/tratamento farmacológico , Receptores Opioides delta/agonistas , Receptores Opioides mu/agonistas , Animais , Ligação Competitiva , Hidromorfona/química , Hidromorfona/farmacologia , Hipercapnia/patologia , Camundongos , Modelos Moleculares , Ligação Proteica , Receptores Opioides delta/antagonistas & inibidores , Receptores Opioides delta/metabolismo , Receptores Opioides mu/antagonistas & inibidores , Receptores Opioides mu/metabolismo , Respiração Artificial , Saimiri , Relação Estrutura-AtividadeRESUMO
Nonribosomal peptide synthetases use tailoring domains to incorporate chemical diversity into the final natural product. A structurally unique set of tailoring domains are found to be stuffed within adenylation domains and have only recently begun to be characterized. PchF is the NRPS termination module in pyochelin biosynthesis and includes a stuffed methyltransferase domain responsible for S-adenosylmethionine (AdoMet)-dependent N-methylation. Recent studies of stuffed methyltransferase domains propose a model in which methylation occurs on amino acids after adenylation and thiolation rather than after condensation to the nascent peptide chain. Herein, we characterize the adenylation and stuffed methyltransferase didomain of PchF through the synthesis and use of substrate analogues, steady-state kinetics, and onium chalcogen effects. We provide evidence that methylation occurs through an SN2 reaction after thiolation, condensation, cyclization, and reduction of the module substrate cysteine and is the penultimate step in pyochelin biosynthesis.
Assuntos
Proteínas de Bactérias/química , Metiltransferases/química , Peptídeo Sintases/química , Fenóis/química , Tiazóis/química , Proteínas de Bactérias/isolamento & purificação , Catálise , Catecol O-Metiltransferase/química , Escherichia coli/genética , Cinética , Methanocaldococcus/enzimologia , Metionina Adenosiltransferase/química , Metionina Adenosiltransferase/isolamento & purificação , Metilação , Metiltransferases/isolamento & purificação , Peptídeo Sintases/isolamento & purificação , Fenóis/síntese química , Domínios Proteicos , Pseudomonas aeruginosa/enzimologia , S-Adenosilmetionina/análogos & derivados , Tiazóis/síntese químicaRESUMO
The kappa (κ) opioid receptor/dynorphin system modulates depression-like states and anhedonia, as well adaptations to stress and exposure to drugs of abuse. Several relatively short-acting small molecule κ-receptor antagonists have been synthesized, and their behavioral profile has been examined under some conditions. The hypothesis of this study is that pharmacological manipulations of the κ-receptor system will result in changes in ethologically relevant anhedonic-like behaviors in mice. Adult male C57BL/6j mice (n = 6-8) were examined for self-grooming behavior in the splash test (in which robust self-grooming is elicited by spraying the dorsum of the mouse with a sucrose solution). The κ-agonist salvinorin A (0.56-1.8 mg/kg) produced dose-dependent decreases in self-grooming, a marker of anhedonia. The selectivity, potency, and duration of action of two relatively short-acting κ-antagonists, LY2444296 [(S)-3-fluoro-4-(4-((2-(3-fluorophenyl) pyrrolidin-1-yl)methyl)phenoxy)benzamide] and LY2795050 [3-chloro-4-(4-(((2S)-2-pyridin-3-ylpyrrolidin-1-yl)methyl) phenoxy)benzamide], were studied for their effectiveness in preventing grooming deficits caused by salvinorin A (1.8 mg/kg). κ-selective doses of both LY2444296 (0.032-1 mg/kg) and LY2795050 (0.032-0.32 mg/kg) dose- and time-dependently prevented the grooming deficits caused by salvinorin A (1.8 m/kg). We also found that a κ-selective dose of each of these antagonists decreased immobility in the forced swim test, a common test of anti-anhedonia effects. This study shows that the κ-receptor system is involved in an ethologically relevant measure of anhedonia, and that κ-selective doses of these antagonists can produce effects consistent with rapid anti-anhedonia. SIGNIFICANCE STATEMENT: Activation of the κ-opioid receptor system results in grooming deficits in mice, an ethologically relevant marker of anhedonia. Shorter acting κ-antagonists are able to cause effects consistent with rapid antianhedonia.
Assuntos
Asseio Animal/efeitos dos fármacos , Antagonistas de Entorpecentes/farmacologia , Receptores Opioides kappa/agonistas , Receptores Opioides kappa/antagonistas & inibidores , Animais , Relação Dose-Resposta a Droga , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nociceptividade/efeitos dos fármacos , Oxicodona/antagonistas & inibidores , Oxicodona/farmacologia , Filosofia , Receptores Opioides kappa/metabolismo , NataçãoRESUMO
Mu opioid receptor (MOR)-targeting analgesics are efficacious pain treatments, but notorious for their abuse potential. In preclinical animal models, coadministration of traditional kappa opioid receptor (KOR)-targeting agonists with MOR-targeting analgesics can decrease reward and potentiate analgesia. However, traditional KOR-targeting agonists are well known for inducing antitherapeutic side effects (psychotomimesis, depression, anxiety, dysphoria). Recent data suggest that some functionally selective, or biased, KOR-targeting agonists might retain the therapeutic effects of KOR activation without inducing undesirable side effects. Nalfurafine, used safely in Japan since 2009 for uremic pruritus, is one such functionally selective KOR-targeting agonist. Here, we quantify the bias of nalfurafine and several other KOR agonists relative to an unbiased reference standard (U50,488) and show that nalfurafine and EOM-salvinorin-B demonstrate marked G protein-signaling bias. While nalfurafine (0.015 mg/kg) and EOM-salvinorin-B (1 mg/kg) produced spinal antinociception equivalent to 5 mg/kg U50,488, only nalfurafine significantly enhanced the supraspinal analgesic effect of 5 mg/kg morphine. In addition, 0.015 mg/kg nalfurafine did not produce significant conditioned place aversion, yet retained the ability to reduce morphine-induced conditioned place preference in C57BL/6J mice. Nalfurafine and EOM-salvinorin-B each produced robust inhibition of both spontaneous and morphine-stimulated locomotor behavior, suggesting a persistence of sedative effects when coadministered with morphine. Taken together, these findings suggest that nalfurafine produces analgesic augmentation, while also reducing opioid-induced reward with less risk of dysphoria. Thus, adjuvant administration of G protein-biased KOR agonists like nalfurafine may be beneficial in enhancing the therapeutic potential of MOR-targeting analgesics, such as morphine.
Assuntos
Analgesia/métodos , Sistemas de Liberação de Medicamentos/métodos , Morfinanos/administração & dosagem , Morfina/administração & dosagem , Medição da Dor/efeitos dos fármacos , Receptores Opioides mu/metabolismo , Compostos de Espiro/administração & dosagem , Animais , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Sinergismo Farmacológico , Feminino , Locomoção/efeitos dos fármacos , Locomoção/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Medição da Dor/métodos , Distribuição Aleatória , Receptores Opioides kappa/administração & dosagem , Receptores Opioides mu/agonistasRESUMO
A reliable protocol to synthesize both racemic and chiral (E)-4-iodobut-3-en-1-ols from aldehydes or epoxides, respectively, containing various aromatic and aliphatic substitutions has been established. The utility of these compounds was then demonstrated by providing access to known fungal decanolides as well as novel aromatic macrocycles. The protocol provided a gram-scale route to (-)-aspinolide A and (-)-5-epi-aspinolide A utilizing a catalytic Nozaki-Hiyama-Kishi reaction to close the macrolide in the final step in 65-84% yields.
Assuntos
Butanóis/síntese química , Lactonas/síntese química , Compostos Macrocíclicos/síntese química , Aldeídos/química , Butanóis/química , Compostos de Epóxi/química , Lactonas/química , Compostos Macrocíclicos/química , Estrutura Molecular , EstereoisomerismoRESUMO
The acute activation of kappa opioid receptors (KOPr) produces antinociceptive and anti-cocaine effects, however, their side-effects have limited further clinical development. Mesyl Sal B is a potent and selective KOPr analogue of Salvinorin A (Sal A), a psychoactive natural product isolated from the plant Salvia divinorum. We assessed the antinociceptive, anti-cocaine, and side-effects of Mesyl Sal B. The anti-cocaine effects are evaluated in cocaine-induced hyperactivity and behavioral sensitization to cocaine in male Sprague Dawley rats. Mesyl Sal B was assessed for anhedonia (conditioned taste aversion), aversion (conditioned place aversion), pro-depressive effects (forced swim test), anxiety (elevated plus maze) and learning and memory deficits (novel object recognition). In male B6.SJL mice, the antinociceptive effects were evaluated in warm-water (50 °C) tail withdrawal and intraplantar formaldehyde (2%) assays and the sedative effects measured with the rotarod performance task. Mesyl Sal B (0.3 mg/kg) attenuated cocaine-induced hyperactivity and behavioral sensitization to cocaine without modulating sucrose self-administration and without producing aversion, sedation, anxiety, or learning and memory impairment in rats. However, increased immobility was observed in the forced swim test indicating pro-depressive effects. Mesyl Sal B was not as potent as Sal A at reducing pain in the antinociceptive assays. In conclusion, Mesyl Sal B possesses anti-cocaine effects, is longer acting in vivo and has fewer side-effects when compared to Sal A, however, the antinociceptive effects are limited.
Assuntos
Comportamento Animal/efeitos dos fármacos , Transtornos Relacionados ao Uso de Cocaína/metabolismo , Transtornos Relacionados ao Uso de Cocaína/psicologia , Cocaína/efeitos adversos , Diterpenos Clerodânicos/farmacologia , Diterpenos/farmacologia , Mesilatos/farmacologia , Receptores Opioides kappa/agonistas , Animais , Ansiedade/tratamento farmacológico , Ansiedade/metabolismo , Transtornos Relacionados ao Uso de Cocaína/tratamento farmacológico , Diterpenos/efeitos adversos , Diterpenos/química , Diterpenos Clerodânicos/efeitos adversos , Diterpenos Clerodânicos/química , Aprendizagem/efeitos dos fármacos , Masculino , Mesilatos/efeitos adversos , Mesilatos/química , Camundongos , Atividade Motora/efeitos dos fármacos , Nociceptividade/efeitos dos fármacos , Dor/tratamento farmacológico , Dor/etiologia , Dor/metabolismo , Ratos , Reconhecimento Psicológico/efeitos dos fármacosRESUMO
In many human alcoholics, abstinence is self-imposed because of the negative consequences of excessive alcohol use, and relapse is often triggered by exposure to environmental contexts associated with prior alcohol drinking. We recently developed a rat model of this human condition in which we train alcohol-preferring P rats to self-administer alcohol in one context (A), punish the alcohol-reinforced responding in a different context (B), and then test for relapse to alcohol seeking in Contexts A and B without alcohol or shock. Here, we studied the role of projections to nucleus accumbens (NAc) shell from ventral subiculum (vSub), basolateral amygdala, paraventricular thalamus, and ventral medial prefrontal cortex in context-induced relapse after punishment-imposed abstinence. First, we measured double-labeling of the neuronal activity marker Fos with the retrograde tracer cholera toxin subunit B (injected in NAc shell) and demonstrated that context-induced relapse is associated with selective activation of the vSubâNAc shell projection. Next, we reversibly inactivated the vSub with GABA receptor agonists (muscimol+baclofen) before the context-induced relapse tests and provided evidence for a causal role of vSub in this relapse. Finally, we used a dual-virus approach to restrict expression of the inhibitory κ opioid-receptor based DREADD (KORD) in vSubâNAc shell projection neurons. We found that systemic injections of the KORD agonist salvinorin B, which selectively inhibits KORD-expressing neurons, decreased context-induced relapse to alcohol seeking. Our results demonstrate a critical role of vSub in context-induced relapse after punishment-imposed abstinence and further suggest a role of the vSubâNAc projection in this relapse. SIGNIFICANCE STATEMENT: In many human alcoholics, abstinence is self-imposed because of the negative consequences of excessive use, and relapse is often triggered by exposure to environmental contexts associated with prior alcohol use. Until recently, an animal model of this human condition did not exist. We developed a rat model of this human condition in which we train alcohol-preferring P rats to self-administer alcohol in one context (A), punish the alcohol-reinforced responding in a different context (B), and test for relapse to alcohol seeking in Contexts A and B. Here, we used neuroanatomical, neuropharmacological, and chemogenetic methods to demonstrate a role of ventral subiculum and potentially its projections to nucleus accumbens in context-induced relapse after punishment-imposed abstinence.