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1.
Life Sci ; 285: 119996, 2021 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-34597607

RESUMO

AIMS: Dezocine and pentazocine, widely prescribed in China for postoperative pain, were initially considered as mixed agonist/antagonist targeting µ-opioid receptors (MORs) and κ-opioid receptors (KORs). However, dezocine has been revealed to alleviate chronic neuropathic pain through MOR activation and norepinephrine reuptake inhibition (NRI). This study investigated dezocine- and pentazocine-induced antinociception and physical dependence development, compared to the typical MOR-NRI opioid tapentadol. MAIN METHODS: Calcium mobilization assay was conducted to assess the potency of the drugs while hot-plate test was performed to compare the antinociception. Physical dependence development was compared with morphine. KEY FINDINGS: Treatment with dezocine, pentazocine and tapentadol stimulated calcium mobilization in HEK293 cells stably expressed MORs but not KORs, whereas dezocine and pentazocine inhibited KOR activities. Subcutaneously injected dezocine-, tapentadol- and pentazocine-induced antinociception dose-dependently, in hot-plate test. Intrathecally injected MOR antagonist CTAP, norepinephrine depletor 6-OHDA and α2-adrenoceptor (α2-AR) antagonist yohimbine partially antagonized dezocine, pentazocine and tapentadol antinociception. Whereas specific KOR antagonist GNTI did not alter their antinociception, the putative inverse KOR agonist nor-BNI reduced dezocine and pentazocine antinociception. Moreover, combined CTAP and 6-OHDA or yohimbine blocked dezocine and tapentadol antinociception but displayed the same partial inhibition on pentazocine antinociception as CTAP alone. Furthermore, compared to morphine and pentazocine, long-term treatment with dezocine and tapentadol produced much less physical dependence-related withdrawal signs, which were restored by spinal 6-OHDA or yohimbine treatment. SIGNIFICANCE: Our findings illustrated that dezocine and tapentadol, but not pentazocine, exert remarkable antinociception in nociceptive pain with less abuse liability via dual mechanisms of MOR activation and NRI.


Assuntos
Analgésicos Opioides/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Dor Nociceptiva/tratamento farmacológico , Pentazocina/farmacologia , Receptores Opioides mu/agonistas , Tapentadol/farmacologia , Tetra-Hidronaftalenos/farmacologia , Inibidores da Captação Adrenérgica/química , Inibidores da Captação Adrenérgica/farmacologia , Analgésicos Opioides/química , Analgésicos Opioides/uso terapêutico , Animais , Compostos Bicíclicos Heterocíclicos com Pontes/química , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Agonismo de Drogas , Antagonismo de Drogas , Células HEK293 , Humanos , Camundongos , Pentazocina/química , Pentazocina/uso terapêutico , Receptores Adrenérgicos/metabolismo , Receptores Opioides kappa/agonistas , Receptores Opioides kappa/antagonistas & inibidores , Receptores Opioides mu/antagonistas & inibidores , Tapentadol/química , Tapentadol/uso terapêutico , Tetra-Hidronaftalenos/química , Tetra-Hidronaftalenos/uso terapêutico
2.
Front Immunol ; 12: 687806, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34326841

RESUMO

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a debilitating multi-systemic chronic condition of unknown aetiology classified as an immune dysfunction syndrome and neurological disorder. The discovery of the widely expressed Transient Receptor Potential Melastatin 3 (TRPM3) as a nociceptor channel substantially targeted by certain opioid receptors, and its implication in calcium (Ca2+)-dependent Natural Killer (NK) cell immune functions has raised the possibility that TRPM3 may be pharmacologically targeted to treat characteristic symptoms of ME/CFS. Naltrexone hydrochloride (NTX) acts as an antagonist to the mu (µ)-opioid receptor thus negating its inhibitory function on TRPM3. Based on the benefits reported by patients on their symptoms, low dose NTX (LDN, 3.0-5.0 mg/day) treatment seems to offer some potential benefit suggesting that its effect may be targeted towards the pathomechanism of ME/CFS. As there is no literature confirming the efficacy of LDN for ME/CFS patients in vitro, this study investigates the potential therapeutic effect of LDN in ME/CFS patients. TRPM3 ion channel activity was measured after modulation with Pregnenolone sulfate (PregS) and ononetin in NK cells on 9 ME/CFS patients taking LDN and 9 age- and sex-matched healthy controls using whole-cell patch-clamp technique. We report that ME/CFS patients taking LDN have restored TRPM3-like ionic currents in NK cells. Small ionic currents with a typical TRPM3-like outward rectification were measured after application of PregS, a TRPM3-agonist, in NK cells from patients taking LDN. Additionally, PregS-evoked ionic currents through TRPM3 were significantly modulated by ononetin, a TRPM3-antagonist, in NK cells from ME/CFS patients taking LDN. These data support the hypothesis that LDN may have potential as a treatment for ME/CFS by characterising the underlying regulatory mechanisms of LDN treatment involving TRPM3 and opioid receptors in NK cells. Finally, this study may serve for the repurpose of marketed drugs, as well as support the approval of prospective randomized clinical studies on the role and dose of NTX in treating ME/CFS patients.


Assuntos
Síndrome de Fadiga Crônica/tratamento farmacológico , Células Matadoras Naturais/efeitos dos fármacos , Naltrexona/administração & dosagem , Antagonistas de Entorpecentes/administração & dosagem , Receptores Opioides mu/antagonistas & inibidores , Canais de Cátion TRPM/efeitos dos fármacos , Adulto , Sinalização do Cálcio/efeitos dos fármacos , Estudos de Casos e Controles , Reposicionamento de Medicamentos , Síndrome de Fadiga Crônica/imunologia , Síndrome de Fadiga Crônica/metabolismo , Síndrome de Fadiga Crônica/fisiopatologia , Feminino , Humanos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Masculino , Potenciais da Membrana/efeitos dos fármacos , Pessoa de Meia-Idade , Naltrexona/efeitos adversos , Antagonistas de Entorpecentes/efeitos adversos , Receptores Opioides mu/metabolismo , Canais de Cátion TRPM/metabolismo , Resultado do Tratamento
3.
Eur J Pharmacol ; 903: 174132, 2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-33933466

RESUMO

Opioid-induced constipation is the most prevalent adverse effect of opioid drugs. Peripherally acting mu opioid receptor antagonists (PAMORAs), including naloxegol, are indicated for the treatment of opioid-induced constipation. The aim of this study was the in vitro and in vivo pharmacological characterization of naloxegol in comparison with naloxone. In vitro experiments were performed to measure calcium mobilization in cells coexpressing opioid receptors and chimeric G proteins and mu receptor interaction with G protein and ß-arrestin 2 using bioluminescence resonance energy transfer. In vivo experiments were performed in mice to measure pain threshold using the tail withdrawal assay and colonic transit using the bead expulsion assay. In vitro, naloxegol behaved as a selective and competitive mu receptor antagonist similarly to naloxone, being 3-10-fold less potent. In vivo, naloxone was effective in blocking fentanyl actions when given subcutaneously (sc), but not per os (po). In contrast, naloxegol elicited very similar effects with sc or po administration counteracting in a dose dependent manner the constipating effects of fentanyl without interfering with the fentanyl mediated analgesia. Thus, a useful PAMORA action could be obtained with naloxegol both after po and sc administration.


Assuntos
Constipação Intestinal/tratamento farmacológico , Morfinanos/farmacologia , Antagonistas de Entorpecentes/farmacologia , Polietilenoglicóis/farmacologia , Administração Oral , Analgésicos Opioides/administração & dosagem , Analgésicos Opioides/efeitos adversos , Analgésicos Opioides/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Células CHO , Cálcio/metabolismo , Constipação Intestinal/induzido quimicamente , Cricetulus , Fentanila/administração & dosagem , Fentanila/efeitos adversos , Fentanila/farmacologia , Injeções Subcutâneas , Masculino , Camundongos , Morfinanos/administração & dosagem , Morfina/farmacologia , Naloxona/administração & dosagem , Naloxona/farmacologia , Antagonistas de Entorpecentes/administração & dosagem , Dor/tratamento farmacológico , Polietilenoglicóis/administração & dosagem , Receptores Opioides mu/antagonistas & inibidores , Receptores Opioides mu/efeitos dos fármacos
4.
Commun Biol ; 4(1): 238, 2021 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-33619305

RESUMO

Antibodies represent powerful tools to examine signal transduction pathways. Here, we present a strategy integrating multiple state-of-the-art methods to produce, validate, and utilize antibodies. Focusing on understudied synaptic proteins, we generated 137 recombinant antibodies. We used yeast display antibody libraries from the B cells of immunized rabbits, followed by FACS sorting under stringent conditions to identify high affinity antibodies. The antibodies were validated by high-throughput functional screening, and genome editing. Next, we explored the temporal dynamics of signaling in single cells. A subset of antibodies targeting opioid receptors were used to examine the effect of treatment with opiates that have played central roles in the worsening of the 'opioid epidemic.' We show that morphine and fentanyl exhibit differential temporal dynamics of receptor phosphorylation. In summary, high-throughput approaches can lead to the identification of antibody-based tools required for an in-depth understanding of the temporal dynamics of opioid signaling.


Assuntos
Anticorpos/farmacologia , Especificidade de Anticorpos , Ensaios de Triagem em Larga Escala , Proteína Quinase C/antagonistas & inibidores , Receptores Opioides mu/antagonistas & inibidores , Sinapses/efeitos dos fármacos , Analgésicos Opioides/farmacologia , Animais , Anticorpos/imunologia , Linhagem Celular Tumoral , Ativação Enzimática , Fentanila/farmacologia , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Morfina/farmacologia , Fosforilação , Proteína Quinase C/imunologia , Proteína Quinase C/metabolismo , Coelhos , Receptores Opioides mu/imunologia , Receptores Opioides mu/metabolismo , Transdução de Sinais , Sinapses/imunologia , Sinapses/metabolismo , Fatores de Tempo
5.
Int J Mol Sci ; 22(2)2021 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-33429857

RESUMO

Opioid peptides and their receptors are expressed in the mammalian retina; however, little is known about how they might affect visual processing. The melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs), which mediate important non-image-forming visual processes such as the pupillary light reflex (PLR), express ß-endorphin-preferring, µ-opioid receptors (MORs). The objective of the present study was to elucidate if opioids, endogenous or exogenous, modulate pupillary light reflex (PLR) via MORs expressed by ipRGCs. MOR-selective agonist [D-Ala2, MePhe4, Gly-ol5]-enkephalin (DAMGO) or antagonist D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) was administered via intravitreal injection. PLR was recorded in response to light stimuli of various intensities. DAMGO eliminated PLR evoked by light with intensities below melanopsin activation threshold but not that evoked by bright blue irradiance that activated melanopsin signaling, although in the latter case, DAMGO markedly slowed pupil constriction. CTAP or genetic ablation of MORs in ipRGCs slightly enhanced dim-light-evoked PLR but not that evoked by a bright blue stimulus. Our results suggest that endogenous opioid signaling in the retina contributes to the regulation of PLR. The slowing of bright light-evoked PLR by DAMGO is consistent with the observation that systemically applied opioids accumulate in the vitreous and that patients receiving chronic opioid treatment have slow PLR.


Assuntos
Peptídeos Opioides/genética , Receptores Opioides mu/genética , Retina/metabolismo , Percepção Visual/genética , Animais , Ala(2)-MePhe(4)-Gly(5)-Encefalina/farmacologia , Encefalinas/antagonistas & inibidores , Encefalinas/genética , Humanos , Luz , Camundongos , Peptídeos/farmacologia , Receptores Opioides/genética , Receptores Opioides mu/antagonistas & inibidores , Reflexo/genética , Retina/patologia , Células Ganglionares da Retina/metabolismo , Células Ganglionares da Retina/fisiologia , Transdução de Sinais/efeitos dos fármacos , Percepção Visual/efeitos dos fármacos , beta-Endorfina/genética
6.
Pharmacogenet Genomics ; 31(4): 75-82, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33395026

RESUMO

In pharmacogenomics, variable receptor phenotypes, resulting from genetic polymorphisms, are often described as a change in protein function or regulation observed upon exposure to a drug. However, in some instances, phenotypes are defined using a class of medications rather than individual drugs. This paradigm assumes that a variation associated with a drug response phenotype will retain the magnitude and direction of the effect for other drugs with the same mechanism of action. However, nonsynonymous polymorphisms may have ligand-specific effects. The purpose of this study was to investigate the potential for point mutations to asymmetrically affect the binding of different drugs to a common target. Ligand binding data from site-directed mutagenesis studies on five G-protein coupled receptors (beta-1 and -2 adrenergic, dopamine D2, angiotensin II and mu-opioid receptor) were collected and analyzed. Binding data from 81 studies for 253 ligands with 447 mutant proteins, including 10 naturally occurring human variants, were analyzed, yielding 1989 mutation-ligand pairs. Fold change in binding affinity for mutant proteins, relative to the wild-type, for different drugs was examined for ligand-specific effects, with a fold-change difference of one or more orders of magnitude between agents considered significant. Of the mutations examined, 49% were associated with ligand-specific effects. One human variant (T164I, beta-2 adrenergic receptor) showed ligand-specific effects for antiasthmatic agents. These results indicate that ligand-specific changes in binding are a possible consequence of missense mutations. This implies that caution needs to be exercised when grouping drugs together during design or interpretation of genotype-phenotype association studies.


Assuntos
Mutagênese Sítio-Dirigida , Testes Farmacogenômicos , Receptores Acoplados a Proteínas G/genética , Receptores Opioides mu/genética , Antagonistas de Receptores de Angiotensina/farmacologia , Estudos de Associação Genética , Humanos , Ligantes , Polimorfismo Genético/genética , Receptores Adrenérgicos beta 1/genética , Receptores Adrenérgicos beta 2/genética , Receptores de Angiotensina/genética , Receptores de Dopamina D2/genética , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Opioides mu/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Mutação Silenciosa/genética
7.
J Ethnopharmacol ; 270: 113872, 2021 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-33485984

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Mu opioid receptor (MOR) is mainly a drug target for analgesia. Opioid-like agonists such as morphine have been clinically used for analgesia but have potential adverse effects. MOR antagonists have been demonstrated to alleviate these side effects. Plants (Carthamus tinctorius L, Cynanchum otophyllum C. K. Schneid., Coffea arabica L., Prinsepia utilis Royle and Lepidium meyenii Walp.) and Ganoderma fungi (Ganoderma hainanense J. D. Zhao, Ganoderma capense (Lloyd) Teng, Ganoderma cochlear (Blume et Nees) Bres., Ganoderma resinaceum Boud and Ganoderma applanatum (Pers.) Pat.) are traditional medicines with beneficial effects on immunoregulation, analgesia and the nervous system, but whether MORs are engaged in their effects remains unknown. AIM OF THE STUDY: This work aimed to identify MOR ligands among compounds isolated from the above-mentioned 10 species, and to investigate selectivity against four opioid receptor subtypes. By analyzing the structure-activity relationship and off-target effects, we could provide a new direction for the future development of MOR drugs. MATERIALS AND METHODS: Four opioid receptor subtype models, including MOR, delta (DOR), kappa (KOR) and nop (NOR), were established with a label-free phenotypic dynamic mass redistribution assay to systematically profile the pharmacological properties of known ligands. Then, 82 natural compounds derived from the 10 species were screened against MOR to identify new ligands. The selectivity of the new ligands was characterized against the four subtypes, and off-target effects were also investigated on eight G protein-coupled receptors (GPCRs). RESULTS: The pharmacological properties of known ligands on transfected HEK293T-MOR, HEK293-DOR, HEK293-KOR and HEK293-NOR cell lines were characterized. Seven compounds purified from Ganoderma cochlear (Blume et Nees) Bres. and Carthamus tinctorius L were MOR antagonists with micromolar potency. Among them, compound 35 showed the strongest antagonistic activity on MOR with an IC50 value of 10.0 ± 3.0 µM. To a certain extent, these seven new antagonists, exhibited antagonistic activity on the other opioid receptor subtypes, and they had almost no effect on other GPCRs, including CB1, CB2, M2 and beta2AR. Additionally, a compound from Lepidium meyenii Walp. displayed MOR agonistic activity. CONCLUSIONS: The established screening models opened new avenues for the discovery and evaluation of opioid receptor ligand selectivity. Together, the novel MOR antagonists and agonists will enrich the inventory of MOR ligands and benefit related therapies.


Assuntos
Produtos Biológicos/farmacologia , Antagonistas de Entorpecentes/farmacologia , Receptores Opioides mu/agonistas , Receptores Opioides mu/antagonistas & inibidores , Receptores Opioides/metabolismo , Produtos Biológicos/química , Técnicas Biossensoriais/métodos , Descoberta de Drogas/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Células HEK293 , Humanos , Ligantes , Receptores Opioides/agonistas , Receptores Opioides/genética , Relação Estrutura-Atividade
8.
J Pharmacol Exp Ther ; 376(3): 374-384, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33443077

RESUMO

Pharmacodynamic efficacy of drugs to activate their receptors is a key determinant of drug effects, and intermediate-efficacy agonists are often useful clinically because they retain sufficient efficacy to produce therapeutically desirable effects while minimizing undesirable effects. Molecular mechanisms of efficacy are not well understood, so rational drug design to control efficacy is not yet possible; however, receptor theory predicts that fixed-proportion mixtures of an agonist and antagonist for a given receptor can be adjusted to precisely control net efficacy of the mixture in activating that receptor. Moreover, the agonist proportion required to produce different effects provides a quantitative scale for comparing efficacy requirements across those effects. To test this hypothesis, the present study evaluated effectiveness of fixed-proportion agonist/antagonist mixtures to produce in vitro and in vivo effects mediated by µ-opioid receptors (MOR) and cannabinoid type 1 receptors (CB1R). Mixtures of 1) the MOR agonist fentanyl and antagonist naltrexone and 2) the CB1R agonist CP55,940 and antagonist/inverse agonist rimonabant were evaluated in an in vitro assay of ligand-stimulated guanosine 5'-O-(3-[35S]thio)triphosphate binding and an in vivo assay of thermal nociception in mice. For both agonist/antagonist pairs in both assays, increasing agonist proportions produced graded increases in maximal mixture effects, and lower agonist proportions were sufficient to produce in vivo than in vitro effects. These findings support the utility of agonist-antagonist mixtures as a strategy to control net efficacy of receptor activation and to quantify and compare efficacy requirements across a range of in vitro and in vivo endpoints. SIGNIFICANCE STATEMENT: Manipulation of agonist proportion in agonist/antagonist mixtures governs net mixture efficacy at the target receptor. Parameters of agonist/antagonist mixture effects can provide a quantitative metric for comparison of efficacy requirements across a wide range of conditions.


Assuntos
Analgésicos Opioides/farmacologia , Canabinoides/farmacologia , Animais , Células CHO , Cricetulus , Relação Dose-Resposta a Droga , Interações Medicamentosas , Masculino , Camundongos , Receptor CB1 de Canabinoide/agonistas , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptores Opioides mu/agonistas , Receptores Opioides mu/antagonistas & inibidores
9.
Psychopharmacology (Berl) ; 238(5): 1255-1263, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-31900526

RESUMO

Functional magnetic resonance imaging (fMRI) has been widely used to gain a greater understanding of brain circuitry abnormalities in CNS disorders. fMRI has also been used to examine pharmacological modulation of brain circuity and is increasingly being used in early clinical drug development as functional pharmacodynamic index of target engagement, and to provide early indication of clinical efficacy. In this short review, we summarize data from experimental medicine and early clinical development studies of a mu-opioid receptor antagonist, GSK1521498 developed for disorders of compulsive consumption including binge eating in obesity. We demonstrate how fMRI can be used to answer important questions of early clinical drug development relating to; (1) target engagement, (2) dose response relationships, (3) differential efficacy and (4) prediction of behavioural and clinically relevant outcomes. We also highlight important methodological factors that need to be considered when conducting fMRI studies in drug development given the challenges faced with small sample sizes in Phase 1 and early proof of mechanism studies. While these data highlight the value of fMRI as a biomarker in drug development, its use for making Go/No-go decisions is still faced with challenges given the variability of responses, interpretation of brain activation changes and the limited data linking drug induced changes in brain activity to clinical or behavioural outcome. These challenges need to be addressed to fulfil the promise of fMRI as a tool in clinical drug development.


Assuntos
Imageamento por Ressonância Magnética/métodos , Antagonistas de Entorpecentes/farmacologia , Receptores Opioides mu/antagonistas & inibidores , Transtorno da Compulsão Alimentar/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Comportamento Compulsivo/tratamento farmacológico , Humanos , Indanos/farmacologia , Obesidade/fisiopatologia , Triazóis/farmacologia
10.
Behav Brain Res ; 397: 112953, 2021 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-33031872

RESUMO

Opioid abuse is a major health problem. The objective of the present study was to evaluate the potentially disruptive side effects and therapeutic potential of a novel antagonist (D24M) of the mu-/delta-opioid receptor (MOR/DOR) heterodimer in male rats. Administration of high doses of D24M (1 & 10 nmol) into the lateral ventricle did not disrupt home cage wheel running. Repeated twice daily administration of increasing doses of morphine (5-20 mg/kg) over 5 days depressed wheel running and induced antinociceptive tolerance measured with the hot plate test. Administration of D24M had no effect on morphine tolerance, but tended to prolong morphine antinociception in non-tolerant rats. Spontaneous morphine withdrawal was evident as a decrease in body weight, a reduction in wheel running and an increase in sleep during the normally active dark phase of the circadian cycle, and an increase in wheel running and wakefulness in the normally inactive light phase. Administration of D24M during the dark phase on the third day of withdrawal had no effect on wheel running. These data provide additional evidence for the clinical relevance of home cage wheel running as a method to assess spontaneous opioid withdrawal in rats. These data also demonstrate that blocking the MOR/DOR heterodimer does not produce disruptive side effects or block the antinociceptive effects of morphine. Although administration of D24M had no effect on morphine withdrawal, additional studies are needed to evaluate withdrawal to continuous morphine administration and other opioids in rats with persistent pain.


Assuntos
Comportamento Animal/efeitos dos fármacos , Morfina/farmacologia , Antagonistas de Entorpecentes/farmacologia , Entorpecentes/farmacologia , Receptores Opioides delta/antagonistas & inibidores , Receptores Opioides mu/antagonistas & inibidores , Sono/efeitos dos fármacos , Síndrome de Abstinência a Substâncias/tratamento farmacológico , Síndrome de Abstinência a Substâncias/fisiopatologia , Animais , Modelos Animais de Doenças , Masculino , Morfina/administração & dosagem , Antagonistas de Entorpecentes/administração & dosagem , Entorpecentes/administração & dosagem , Ratos , Ratos Sprague-Dawley , Corrida
11.
J Cancer Res Clin Oncol ; 147(3): 779-792, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33315125

RESUMO

PURPOSE: D,L-methadone (MET), an analgesic drug used for pain treatment and opiate addiction, has achieved attention from oncologists and social media as possible chemoensitizing agent in cancer therapy, notably brain cancer (glioblastoma multiforme, GBM). MET has been reported to enhance doxorubicin-induced cytotoxicity in GBM cells via activation of the µ-opioid receptor (MOR). Here, we extended this work and quantified the toxic effect of MET in comparison to other opioids alone and in combination with doxorubicin and the clinically more relevant alkylating drug temozolomide (TMZ), using a set of GBM cell lines and primary GBM cells. METHODS: MOR expression in GBM cells was investigated by immunofluorescence and immunoblotting. Resistance to drugs alone and in combination with anticancer drugs was assessed by MTT assays. Concentration effect curves were fitted by nonlinear regression analysis and IC50 values were calculated. Apoptosis and necrosis rates were determined by annexin V/propidium iodide (PI)-flow cytometry. RESULTS: MET alone was cytotoxic in all GBM cell lines and primary GBM cells at high micromolar concentrations (IC50 ~ 60-130 µM), observed both in the metabolic MTT assay and by quantifying apoptosis and necrosis, while morphine and oxycodone were not cytotoxic in this concentration range. Naloxone was not able to block MET-induced cytotoxicity, indicating that cell death-inducing effects of MET are not MOR-dependent. We recorded doxorubicin and TMZ concentration- response curves in combination with fixed MET concentrations. MET enhanced doxorubicin-induced cytotoxicity in only one cell line, and in primary cells it was observed only in a particular MET concentration range. In all assays, MET was not effective in sensitizing cells to TMZ. In two cell lines, MET even decreased the cell's sensitivity to TMZ. CONCLUSION: MET was found to be cytotoxic in GBM cells in vitro only at high, clinically not relevant concentrations, where it was effective in inducing apoptosis and necrosis. Sensitizing effects were only observed in combination with doxorubicin, but not with TMZ, and are dependent on cell line and the applied drug concentration. Therefore, our findings do not support the use of MET in the treatment of GBM in combination with TMZ, as no sensitizing effect of MET was observed.


Assuntos
Analgésicos Opioides/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias Encefálicas/tratamento farmacológico , Doxorrubicina/farmacologia , Glioblastoma/tratamento farmacológico , Metadona/farmacologia , Analgésicos Opioides/administração & dosagem , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Doxorrubicina/administração & dosagem , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Metadona/administração & dosagem , Morfina/farmacologia , Naloxona/farmacologia , Oxicodona/farmacologia , Receptores Opioides mu/agonistas , Receptores Opioides mu/antagonistas & inibidores , Receptores Opioides mu/biossíntese , Células Tumorais Cultivadas
12.
Int J Mol Sci ; 21(23)2020 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-33255594

RESUMO

The opioid-induced analgesia is associated with a number of side effects such as addiction, tolerance and respiratory depression. The involvement of neuropeptide FF (NPFF) in modulation of pain perception, opioid-induced tolerance and dependence was well documented in contrast to respiratory depression. Therefore, the aim of the present study was to examine the potency of NPFF to block post-opioid respiratory depression, one of the main adverse effects of opioid therapy. Urethane-chloralose anaesthetized Wistar rats were injected either intravenously (iv) or intracerebroventricularly (icv) with various doses of NPFF prior to iv endomorphin-1 (EM-1) administration. Iv NPFF diminished the number of EM-1-induced apneas without affecting their length and without influence on the EM-1 induced blood pressure decline. Icv pretreatment with NPFF abolished the occurrence of post-EM-1 apneas and reduced also the maximal drop in blood pressure and heart rate. These effects were completely blocked by the NPFF receptor antagonist RF9, which was given as a mixture with NPFF before systemic EM-1 administration. In conclusion, our results showed that centrally administered neuropeptide FF is effective in preventing apnea evoked by stimulation of µ-opioid receptors and the effect was due to activation of central NPFF receptors. Our finding indicates a potential target for reversal of opioid-induced respiratory depression.


Assuntos
Apneia/tratamento farmacológico , Doenças Cardiovasculares/tratamento farmacológico , Oligopeptídeos/farmacologia , Receptores Opioides mu/genética , Analgesia/efeitos adversos , Analgésicos Opioides/efeitos adversos , Animais , Apneia/induzido quimicamente , Apneia/genética , Apneia/patologia , Doenças Cardiovasculares/induzido quimicamente , Doenças Cardiovasculares/genética , Modelos Animais de Doenças , Humanos , Infusões Intraventriculares , Oligopeptídeos/efeitos adversos , Oligopeptídeos/genética , Percepção da Dor/efeitos dos fármacos , Ratos , Receptores de Neuropeptídeos/antagonistas & inibidores , Receptores de Neuropeptídeos/genética , Receptores Opioides mu/antagonistas & inibidores , Ativação Transcricional/efeitos dos fármacos
13.
Mol Brain ; 13(1): 150, 2020 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-33176836

RESUMO

The Sigma-1 receptor (σ1R) has emerged as an interesting pharmacological target because it inhibits analgesia mediated by mu-opioid receptors (MOR), and also facilitates the development of neuropathic pain. Based on these findings, the recent cloning of the Sigma-2 receptor (σ2R) led us to investigate its potential role as a regulator of opioid analgesia and of pain hypersensitivity in σ2R knockout mice. In contrast to σ1R deficient mice, σ2R knockout mice developed mechanical allodynia following establishment of chronic constriction injury-induced neuropathic pain, which was alleviated by the σ1R antagonist S1RA. The analgesic effects of morphine, [D-Ala, N-MePhe, Gly-ol]-encephalin (DAMGO) and ß-endorphin increased in σ1R-/- mice and diminished in σ2R-/- mice. The analgesic effect of morphine was increased in σ2R-/- mice by treatment with S1RA. However, σ2R-/- mice and wild-type mice exhibited comparable antinociceptive responses to the delta receptor agonist [D-Pen2,5]-encephalin (DPDPE), the cannabinoid type 1 receptor agonist WIN55,212-2 and the α2-adrenergic receptor agonist clonidine. Therefore, while σR1 inhibits and σ2R facilitates MOR-mediated analgesia these receptors exchange their roles when regulating neuropathic pain perception. Our study may help identify new pharmacological targets for diminishing pain perception and improving opioid detoxification therapies.


Assuntos
Nociceptividade , Receptores Opioides mu/antagonistas & inibidores , Receptores sigma/metabolismo , Analgésicos/farmacologia , Animais , Constrição Patológica , Hiperalgesia/metabolismo , Hiperalgesia/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Morfina/farmacologia , Nociceptividade/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Opioides mu/metabolismo
14.
Mol Pain ; 16: 1744806920966144, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33108956

RESUMO

Lots of studies have demonstrated that anterior cingulate cortex plays important roles in the pain perception and pain modulation. The present study explored the role of mu-opioid receptor in nociceptive modulation in anterior cingulate cortex of rats with neuropathic pain. Neuropathic pain model was set up by chronic constriction injury of the left sciatic nerve of rats. The hindpaw withdrawal latency to thermal and mechanical stimulation, by hot plate and Randall Selitto Test respectively, was used to evaluate the rat's responses to noxious stimulation. Results showed that intra-anterior cingulate cortex injection of morphine could induce the antinociception dose-dependently. By intra-anterior cingulate cortex injection of opioid receptor antagonist, the morphine-induced antinociception could be attenuated by naloxone, as well as much significantly by the selective mu-opioid receptor antagonist ß-funaltrexamine, indicating that mu-opioid receptor is involved in the morphine-induced antinociception in anterior cingulate cortex of rats with neuropathic pain. The morphine-induced antinociception was much more decreased in rats with neuropathic pain than that in normal rats, and there was a significant decrease in mu-opioid receptor messenger RNA levels in anterior cingulate cortex of rats with neuropathic pain, indicating that there may be a down-regulation in mu-opioid receptor expression in anterior cingulate cortex of rats with neuropathic pain. To further confirm the role of mu-opioid receptor in morphine-induced antinociception in anterior cingulate cortex, normal rats were received intra-anterior cingulate cortex administration of small interfering RNA targeting mu-opioid receptor and it was found that there was a down-regulation in mu-opioid receptor messenger RNA levels, as well as a down-regulation in mu-opioid receptor expression in anterior cingulate cortex tested by real-time polymerase chain reaction and western blotting. Furthermore, the morphine-induced antinociceptive effect decreased significantly in rats with small interfering RNA targeting mu-opioid receptor, which indicated that knockdown mu-opioid receptor in anterior cingulate cortex could also attenuate morphine-induced antinociceptive effect. These results strongly suggest that mu-opioid receptor plays a significant role in nociceptive modulation in anterior cingulate cortex of rats.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Giro do Cíngulo/metabolismo , Neuralgia/metabolismo , Nociceptividade , Receptores Opioides mu/antagonistas & inibidores , Receptores Opioides mu/metabolismo , Animais , Escala de Avaliação Comportamental , Regulação da Expressão Gênica/genética , Técnicas de Silenciamento de Genes , Masculino , Morfina/farmacologia , Naloxona/farmacologia , Naltrexona/análogos & derivados , Naltrexona/farmacologia , RNA Interferente Pequeno , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Receptores Opioides mu/genética , Nervo Isquiático/lesões
15.
Molecules ; 25(19)2020 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-33007971

RESUMO

Increasing evidence indicates that native mu and delta opioid receptors can associate to form heteromers in discrete brain neuronal circuits. However, little is known about their signaling and trafficking. Using double-fluorescent knock-in mice, we investigated the impact of neuronal co-expression on the internalization profile of mu and delta opioid receptors in primary hippocampal cultures. We established ligand selective mu-delta co-internalization upon activation by 1-[[4-(acetylamino)phenyl]methyl]-4-(2-phenylethyl)-4-piperidinecarboxylic acid, ethyl ester (CYM51010), [d-Ala2, NMe-Phe4, Gly-ol5]enkephalin (DAMGO), and deltorphin II, but not (+)-4-[(αR)-α-((2S,5R)-4-Allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80), morphine, or methadone. Co-internalization was driven by the delta opioid receptor, required an active conformation of both receptors, and led to sorting to the lysosomal compartment. Altogether, our data indicate that mu-delta co-expression, likely through heteromerization, alters the intracellular fate of the mu opioid receptor, which provides a way to fine-tune mu opioid receptor signaling. It also represents an interesting emerging concept for the development of novel therapeutic drugs and strategies.


Assuntos
Receptores Opioides delta/antagonistas & inibidores , Receptores Opioides mu/antagonistas & inibidores , Animais , Células Cultivadas , Endocitose , Hipocampo/citologia , Ligantes , Lisossomos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Piperidinas/farmacologia , Multimerização Proteica , Receptores Opioides delta/metabolismo , Receptores Opioides mu/metabolismo
16.
Curr Top Med Chem ; 20(31): 2866-2877, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32962616

RESUMO

Since a µ-opioid receptor gene containing multiple exons has been identified, the variety of splice variants for µ-opioid receptors have been reported in various species. Amidino-TAPA and IBNtxA have been discovered as new analgesics with different pharmacological profiles from morphine. These new analgesics show a very potent analgesic effect but do not have dependence liability. Interestingly, these analgesics show the selectivity to the morphine-insensitive µ-opioid receptor splice variants. The splice variants, sensitive to these new analgesics but insensitive to morphine, may be a better molecular target to develop the analgesics without side effects.


Assuntos
Analgésicos Opioides/farmacologia , Receptores Opioides mu/antagonistas & inibidores , Processamento Alternativo/efeitos dos fármacos , Processamento Alternativo/genética , Analgésicos Opioides/química , Humanos , Receptores Opioides mu/genética
17.
Peptides ; 134: 170407, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32926948

RESUMO

The present study was undertaken to further investigate the spinal anti-allodynic effects of endomorphins (EMs) and their C-terminal hydrazide modified analogs EM-1-NHNH2 and EM-2-NHNH2 in the spared nerve injury (SNI) model of neuropathic pain in mice. Our results demonstrated that intrathecal (i.t.) administration of endomorphin-1 (EM-1), endomorphin-2 (EM-2), EM-1-NHNH2 and EM-2-NHNH2 produced potent anti-allodynic effects ipsilaterally in neuropathic pain model. Judging from the area under the curve (AUC) values, these two analogs exhibited higher antinociception than their parent peptides. Moreover, they also displayed significant antinociceptive effects in the contralateral paw administered intrathecally. Interestingly, EM-1 and its analog EM-1-NHNH2 displayed their antinociception probably by µ2-opioid receptor subtype since the µ1-opioid receptor antagonist naloxonazine didn't significantly block the anti-allodynia of EM-1 and EM-1-NHNH2, which implied a same opioid mechanism. However, the anti-allodynia induced by EM-2, but not EM-2-NHNH2 was significantly reduced by both µ1-opioid antagonist, naloxonazine and κ-antagonist, nor-binaltorphamine (nor-BNI), indicating multiple opioid receptors were involved in the anti-allodynic effects of EM-2. Most importantly, EM-1-NHNH2 decreased the antinociceptive tolerance, and EM-2-NHNH2 displayed non-tolerance-forming antinociception. Therefore, C-terminal amide to hydrazide conversion changed the spinal antinociceptive profiles of EMs in neuropathic pain. The present investigation is of great value in the development of novel opioid therapeutics against neuropathic pain.


Assuntos
Azidas/química , Hiperalgesia/tratamento farmacológico , Neuralgia/tratamento farmacológico , Oligopeptídeos/farmacologia , Receptores Opioides mu/metabolismo , Analgésicos Opioides/farmacologia , Animais , Modelos Animais de Doenças , Hiperalgesia/metabolismo , Hiperalgesia/patologia , Injeções Espinhais/métodos , Masculino , Camundongos , Antagonistas de Entorpecentes/farmacologia , Neuralgia/metabolismo , Neuralgia/patologia , Oligopeptídeos/química , Medição da Dor/métodos , Receptores Opioides mu/antagonistas & inibidores
18.
Future Med Chem ; 12(22): 2001-2018, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32972243

RESUMO

Background: Central and peripheral analgesia without adverse effects relies on the identification of µ-opioid agonists that are able to activate 'basal' antinociceptive pathways. Recently developed µ-selective benzomorphan agonists that are not antagonized by naloxone do not activate G-proteins and ß-arrestins. Which pathways do µ receptors activate? How can each of them be selectively activated? What role is played by allosteric binding sites? Methodology & results: Molecular modeling studies characterize the amino acid residues involved in the interaction with various classes of endogenous and exogenous ligands and with agonists and antagonists. Conclusions: Critical binding differences between various classes of agonists with different pharmacological profiles have been identified. MML series binding poses may be relevant in the search for an antinociception agent without side effects.


Assuntos
Analgésicos Opioides/farmacologia , Simulação de Dinâmica Molecular , Receptores Opioides mu/agonistas , Receptores Opioides mu/antagonistas & inibidores , Analgésicos Opioides/química , Sítios de Ligação/efeitos dos fármacos , Humanos , Ligantes , Estrutura Molecular
19.
Psychopharmacology (Berl) ; 237(10): 3057-3065, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32772146

RESUMO

RATIONALE: Opioid abuse remains a serious public health problem. The pseudoirreversible mu opioid receptor antagonist methocinnamox (MCAM) might be useful for treating opioid abuse and overdose. Because endogenous opioid systems can modulate cognition and decision-making, it is important to evaluate whether long-term blockade of mu opioid receptors by MCAM adversely impacts complex operant behavior involving memory. OBJECTIVE: This study tested the effects of MCAM in rhesus monkeys responding under a delayed matching-to-sample task, with correct responses reinforced by sucrose pellets. Because MCAM did not alter performance, antagonism of the rate-decreasing effects of morphine was used to confirm that an effective dose of MCAM was administered. Moreover, the muscarinic receptor antagonist scopolamine and the N-methyl-D-aspartate antagonist phencyclidine were studied as positive controls to demonstrate sensitivity of this procedure to memory disruption. RESULTS: Neither MCAM (0.32 mg/kg) nor morphine (1-5.6 mg/kg) impaired delayed matching-to-sample accuracy. Morphine dose-dependently decreased the number of trials completed before MCAM administration, and a single injection of MCAM blocked the behavioral suppressant effects of morphine for at least 7 days. Scopolamine (0.01-0.056 mg/kg) and phencyclidine (0.1-0.56 mg/kg) dose-dependently decreased delayed matching-to-sample accuracy and the number of trials completed. CONCLUSIONS: MCAM did not impair memory (as measured by accuracy in a delayed matching-to-sample task) and did not decrease responding for or consumption of sucrose pellets. This dose of MCAM attenuates self-administration of opioids and reverses as well as prevents opioid-induced respiratory depression. These results provide further support for a favorable adverse effect profile for MCAM.


Assuntos
Cinamatos/farmacologia , Memória/efeitos dos fármacos , Derivados da Morfina/farmacologia , Morfina/antagonistas & inibidores , Morfina/farmacologia , Antagonistas de Entorpecentes/farmacologia , Receptores Opioides mu/antagonistas & inibidores , Animais , Cognição/efeitos dos fármacos , Cognição/fisiologia , Relação Dose-Resposta a Droga , Feminino , Macaca mulatta , Masculino , Memória/fisiologia , Receptores Opioides mu/fisiologia , Reforço Psicológico , Autoadministração
20.
Am J Clin Dermatol ; 21(5): 601-618, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32607945

RESUMO

Chronic pruritus, defined as an unpleasant sensation resulting in a need to scratch that lasts more than 6 weeks, is a prevalent and bothersome symptom associated with both cutaneous and systemic conditions. Due to complex pathogenesis and profuse contributing factors, chronic pruritus therapy remains challenging. Regardless of the well-established antipruritic properties of classic pharmacotherapy (topical therapy, phototherapy and systemic therapy), these methods often provide insufficient relief for affected individuals. Owing to the growing interest in the field of pruritic research, further experimental and clinical data have emerged, continuously supporting the possibility of instigating novel therapeutic measures. This review covers the most relevant current modalities remaining under investigation that possess promising perspectives of approval in the near future, especially opioidergic drugs (mu-opioid antagonists and kappa-opioid agonists), neurokinin-1 receptor antagonists, biologic drugs, Janus kinase inhibitors, ileal bile acid transporter inhibitors, aryl hydrocarbon receptor agonists and histamine H4 receptor antagonists.


Assuntos
Prurido/tratamento farmacológico , Analgésicos Opioides/farmacologia , Analgésicos Opioides/uso terapêutico , Antipruriginosos/uso terapêutico , Fatores de Transcrição Hélice-Alça-Hélice Básicos/agonistas , Fatores Biológicos/uso terapêutico , Doença Crônica/terapia , Ensaios Clínicos como Assunto , Humanos , Inibidores de Janus Quinases/uso terapêutico , Antagonistas dos Receptores de Neurocinina-1/uso terapêutico , Fototerapia/métodos , Prurido/etiologia , Prurido/psicologia , Qualidade de Vida , Receptores de Hidrocarboneto Arílico/agonistas , Receptores Histamínicos H4/antagonistas & inibidores , Receptores Opioides kappa/agonistas , Receptores Opioides mu/antagonistas & inibidores , Pele/inervação , Pele/patologia , Resultado do Tratamento
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