Morphine Metabolites Research Articles

Physiologically Based Pharmacokinetic (PBPK) Model Predictions of Disease Mediated Changes in Drug Disposition in Patients with Nonalcoholic Fatty Liver Disease (NAFLD).

This study was designed to verify a virtual population representing patients with nonalcoholic fatty liver disease (NAFLD) to support the implementation of a physiologically based pharmacokinetic (PBPK) modeling approach for prediction of disease-related changes in drug pharmacokinetics. A virtual NAFLD patient population was developed in GastroPlus (v.9.8.2) by accounting for pathophysiological changes associated with the disease and proteomics-informed alterations in the abundance of metabolizing enzymes and transporters pertinent to drug disposition. The NAFLD population model was verified using exemplar drugs where elimination is influenced predominantly by cytochrome P450 (CYP) enzymes (chlorzoxazone, caffeine, midazolam, pioglitazone) or by transporters (rosuvastatin, 11C-metformin, morphine and the glucuronide metabolite of morphine). PBPK model predictions of plasma concentrations of all the selected drugs and hepatic radioactivity levels of 11C-metformin were consistent with the clinically-observed data. Importantly, the PBPK simulations using the virtual NAFLD population model provided reliable estimates of the extent of changes in key pharmacokinetic parameters for the exemplar drugs, with mean predicted ratios (NAFLD patients divided by healthy individuals) within 0.80- to 1.25-fold of the clinically-reported values, except for midazolam (prediction-fold difference of 0.72). A virtual NAFLD population model within the PBPK framework was successfully developed with good predictive capability of estimating disease-related changes in drug pharmacokinetics. This supports the use of a PBPK modeling approach for prediction of the pharmacokinetics of new investigational or repurposed drugs in patients with NAFLD and may help inform dose adjustments for drugs commonly used to treat comorbidities in this patient population.

Activation of Akt–cAMP response element-binding protein (CREB) signaling as an adaptive response to an electrophilic metabolite of morphine

Activation of Akt–cAMP response element-binding protein (CREB) signaling as an adaptive response to an electrophilic metabolite of morphine

Blocking Palmitoylation of Apelin Receptor Alleviates Morphine Tolerance in Neuropathic Cancer Pain.

Neuropathic cancer pain (NCP) is an important symptom in patients with cancer. However, significant analgesic tolerance and other side effects critically hamper the administration of morphine. Protein palmitoylation mediated by the DHHC family may be involved in the glial activation and inflammatory responses underlying organ failure. In this study, we investigated the key role of protein palmitoylation in cancer pain and sought to target palmitoylation to suppress morphine tolerance. We found that long-term use of morphine led to the accumulation of the morphine metabolite, morphine-3-glucuronide, in vivo and activated ERK1/2 and microglia to release inflammatory factors through the apelin receptor APLNR. Palmitoyltransferase ZDHHC9 was upregulated in NCP, and APLNR was palmitylated to protect it from lysosomal degradation and to maintain its stability. We also designed competitive inhibitors of APLNR palmitoylation to inhibit the development of NCP, release of inflammatory factors, and attenuation of morphine tolerance. Therefore, targeting APLNR palmitoylation in combination with morphine is a potent method for cancer pain treatment. Our data provide a basis for the future clinical use of related drugs combined with morphine for the treatment of cancer-related pain.

Comparative Toxicology of Syringe Exchange and Post-Mortem Blood Samples in the District of Columbia: Trends and Affinity Analysis.

This cross-sectional analysis aimed to understand the similarities and differences between drugs detected in syringes collected from syringe service providers in the District of Columbia and fatal overdose deaths captured by the State Unintentional Drug Overdose Reporting System. Substance exposures for these fatal and non-fatal drug use outcomes have not been previously compared. Substance distributions were examined and a paired significance test was used to compare changes over time. Affinity analysis was employed to reveal substance co-occurrences. Between September, 2020-2022, 1,118 post-mortem blood samples and 3,646 syringes were processed, with fatal overdoses increasing 24.1%. Polysubstance use was more commonly found in post-mortem blood (82.5%) than syringe exchange samples (48.6%). Of samples containing opioids, 94.8% of blood samples and 86.3% of syringes contained fentanyl, fentanyl analogs, or fentanyl precursors/metabolites. Post-mortem blood samples had double the frequency of co-occurring stimulants and opioids (43.9%) as syringe exchange samples (21.8%). Major changes in occurrence frequency over time were found for opioid and stimulant exposure in both groups, especially in the increased occurrence of fluorofentanyl (>400%), methamphetamine (>90%), and xylazine (>60%) while the incidence of fentanyl, heroin, and metabolite morphine declined. These results indicate that while fatal use and syringe exchange populations have distinct substance exposures which may contribute to differences in mortality rate, their substance distributions have similar change magnitudes. This study highlights the utility of using multiple data sources to provide a comprehensive description of drug use patterns and discusses the limitations in reporting data from each source.

Concentrations, pharmacokinetics and selected pharmacodynamics of morphine and its active metabolites following oral administration to horses.

The metabolism and pharmacokinetics of intravenous (i.v.) morphine in the horse have been described; however, administration of therapeutic doses has also been associated with neuroexcitation and adverse gastrointestinal effects. In this study, we hypothesized that oral administration would lead to comparable concentrations of morphine and its presumed active metabolite, morphine 6-glucuronide (M6G) without the adverse effects associated with i.v. administration. Eight horses were administered a single i.v. dose of 0.2mg/kg morphine and oral doses of 0.2, 0.6, and 0.8mg/kg of morphine in a four-way balanced crossover design, with a 2-week washout period between doses. Concentrations of morphine and metabolites were determined, and pharmacokinetic parameters determined. Physiologic and behavioral outcomes including the number of steps taken, changes in heart rate, and gastrointestinal borborygmi were assessed. Oral administration of morphine resulted in higher concentrations of morphine metabolites, including M6G (Cmax : 11.6-37.8ng/mL (0.6mg/kg); 15.8-42.6ng/mL (0.8mg/kg)), compared with i.v. Bioavailability was 36.5%, 27.6% and 28.0% for 0.2, 0.6 and 0.8mg/kg, respectively. Behavioral and physiologic changes were noted in all groups but were less prominent with oral compared with i.v. administration. Results of the current study are encouraging for further study, specifically anti-nociceptive effects of morphine following oral administration.

Activation of the Keap1/Nrf2 Pathway as an Adaptive Response to an Electrophilic Metabolite of Morphine.

Morphinone (MO) is an electrophilic metabolite of morphine that covalently binds to protein thiols via its α,β-unsaturated carbonyl group, resulting in toxicity in vitro and in vivo. Our previous studies identified a variety of redox signaling pathways that are activated during electrophilic stress. Here, we examined in vitro activation of a signaling pathway involving Kelch-like ECH-associated protein 1 (Keap1) and nuclear factor erythroid 2-related factor 2 (Nrf2) in response to MO. Exposure of HepG2 cells to MO caused covalent modification of Keap1 thiols (evaluated using biotin-PEAC5-maleimide labeling) and nuclear translocation of Nrf2, thereby up-regulating downstream genes encoding ATP binding cassette subfamily C member 2, solute carrier family 7 member 11, glutamate-cysteine ligase catalytic subunit, glutamate-cysteine ligase modifier subunit, glutathione S-transferase alpha 1, and heme oxygenase 1. However, dihydromorphinone, a metabolite of morphine lacking the reactive C7-C8 double bond, had little effect on Nrf2 activation. These results suggest that covalent modification is crucial in the Keap1/Nrf2 pathway activation and that this pathway is a redox signaling-associated adaptive response to MO metabolism.

Activation of HSP90/HSF1 Signaling as an Adaptive Response to an Electrophilic Metabolite of Morphine.

Morphinone (MO) is an electrophilic metabolite of morphine that covalently binds to protein thiols, resulting in toxicity in vitro and in vivo. We have previously identified a variety of redox signaling pathways that are activated during electrophilic stress. However, the role of MO in such activation remains unknown. In this study, we examined whether MO could activate heat shock protein (HSP) 90/heat shock factor (HSF) 1 signaling in HepG2 cells. MO exposure caused S-modification of HSP90 (determined using biotin-PEAC5-maleimide labeling) and nuclear translocation of transcription factor HSF1, thereby up-regulating its downstream genes encoding B-cell lymphoma 2-associated anthanogene 3 and heat shock 70 kDa protein 1. However, dihydromorphinone, a non-electrophilic metabolite of morphine, had little effect on HSF1 activation or upregulation of these genes, suggesting that covalent modification plays a role in this process and that the HSP90/HSF1 pathway is a redox-signaled adaptive response to morphine metabolism.

Dissecting the innate immune recognition of morphine and its metabolites by TLR4/MD2: an in silico simulation study.

A toll-like receptor 4/myeloid differentiation factor 2 complex (TLR4/MD2) has been identified as a non-classical opioid receptor capable of recognizing morphine isomers and activating microglia in a non-enantioselective manner. Additionally, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G), the major metabolites of morphine, possess similar chemical structures but exhibit distinct effects on TLR4 signaling. However, the specific mechanisms by which morphine isomers and morphine metabolites are recognized by the innate immune receptor TLR4/MD2 are not well understood. Herein, molecular dynamics simulations were performed to dissect the molecular recognition of TLR4/MD2 with morphine isomers, M3G and M6G. Morphine and its (+)-enantiomer, dextro-morphine ((+)-morphine), were found to have comparable binding free energies as well as similar interaction modes when interacting with (TLR4/MD2)2. Binding with morphine and (+)-morphine caused the motion of the F126 loop towards the inside of the MD2 cavity, which stabilizes (TLR4/MD2)2 with similar dimerization interfaces. The binding free energies of M3G and M6G with (TLR4/MD2)2, while lower than those of morphine isomers, were comparable to each other. However, the binding behaviors of M3G and M6G exhibited contrasting patterns when interacting with (TLR4/MD2)2. The glucuronide group of M3G bound to the gating loop of MD2 and formed strong interactions with TLR4*, which stabilizes the active heterotetrameric complex. In contrast, M6G was situated in cavity A of MD2, where the critical interactions between M6G and the residues of TLR4* were lost, resulting in fluctuation of (TLR4/MD2)2 away from the active conformation. These results indicate that the pivotal interactions at the dimerization interface between MD2 and TLR4* in M6G-bound (TLR4/MD2)2 were considerably weaker than those in M3G-bound (TLR4/MD2)2, which partially explains why M6G fails to activate TLR4 signaling. The discoveries from this study will offer valuable insights for the advancement of next-generation TLR4 small molecule modulators based on opioids.

Pharmacokinetics and transplacental transfer of codeine and codeine metabolites from Papaver somniferum L.

Ethnopharmacological relevancePapaveris Pericarpium, which is the dried husk of Papaver somniferum L., has been used as a phytomedicine to relieve cough, diarrhea and pain. The alkaloid codeine contained therein via biotransformation converts to morphine and potentially produces addictive and toxic effects. Due to the healthy concern for a pregnant woman, our hypothesis is that codeine and its metabolites can penetrate the placental barrier to reach the foetus and amniotic fluid, and these processes may be modulated by the transporter. Aim of the studyBecause codeine is also considered a prodrug of morphine, it has a good analgesic effect. It is often used by pregnant women but may expose the foetus to the risk of morphine harm. The aim of this study is to investigate the metabolic rate, distribution and transplacental transfer mechanism of codeine and its metabolites morphine and morphine-3-glucuronide (M3G) in pregnant rats and to assess the risk of medication for pregnant women. Materials and methodsUltrahigh-performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) combined with a microdialysis system was developed to monitor codeine, morphine and M3G in multiple sites of maternal blood, placenta, foetus and amniotic fluid after codeine administration. A compartmental model was used to calculate the pharmacokinetic parameters of codeine in blood after codeine administration (10 mg/kg, i.v.). The area under the concentration (AUC) ratio of AUCmetabolite/AUCcodeine and AUCtissue/AUCblood was used to represent the metabolic biotransformation ratio and the drug from blood-to-tissue transfer ratio, respectively. ResultsThe pharmacokinetic results demonstrated that codeine fit well with a two-compartment model and went through rapid metabolism to morphine and M3G in pregnant rats after codeine administration (10 mg/kg, i.v.). The biotransformation ratios of AUCmorphine/AUCcodeine, AUCM3G/AUCmorphine and AUCM3G/AUCcodeine were 0.12 ± 0.03, 54.45 ± 20.61 and 6.53 ± 2.47, respectively, after codeine administration (10 mg/kg, i.v.), which suggested that codeine was easily metabolized into M3G through morphine. The tissue distribution results demonstrated that all of the analytes penetrated into the foetus through the placenta; however, the blood-to-tissue transfer ratio (AUCtissue/AUCblood) of morphine and M3G was relatively lower than that of codeine after codeine administration (10 mg/kg, i.v.), which suggested that the blood-placenta barrier blocks the penetration of morphine and M3G into the foetus. Thus, the tissue transfer of morphine in the placenta and foetus was significantly enhanced by treatment with corticosterone, an inhibitor of organic cation transporter (OCT). ConclusionBased on microdialysis coupled to a validated UHPLC–MS/MS system, the pharmacokinetics and metabolic biotransformation of codeine and its metabolites were analyzed and clarified. The potential mechanism of morphine placental transfer was modulated by OCT transporters.

Extrapyramidal syndrome in a breastfed child, relevance of toxicological analysis

The passage of drugs in breast milk may be responsible for toxicity in breastfed children. We report here a case of passage of metoclopramide, an antiemetic neuroleptic, in breast milk, causing an extrapyramidal syndrome in the child, revealed in blood and hair. Case: a 21-week-old child was brought by his parents to the pediatric neurology department for consultation following two episodes of unexplained extrapyramidal syndrome. The little G., fed by mixed breastfeeding, presented a first episode of unexplained acute dystonia (generalized hypertonia with legs in extension and arms flexed against the chest, ceiling gaze, opisthonos) and two weeks later, a new episode of acute extrapyramidal syndrome, which resolved in a few hours. Both events required emergency hospitalization. During the second episode, diazepam 0.5 mg/kg IV was initiated, unsuccessfully. The anamnesis revealed that the mother had taken a rectal suppository of metoclopramide (10 mg) 48 hours before the first episode. An MRI was performed on arrival at the emergency room. Blood toxicology analysis was performed two days after the first episode. A 3-cm strand of brown hair was collected one month after the second episode. The entire sample (13 mg) was analyzed without segmentation. It should also be noted that the father had retractile capsulitis, treated with codeine paracetamol, and that the child had received paracetamol for teething. The child had no further episodes of dystonia since the second event. The MRI was normal. The child's blood toxicology analysis revealed the presence of metoclopramide at a concentration of 8 ng/mL. Hair analysis revealed metoclopramide at a relatively high concentration (1058 pg/mg), as well as paracetamol (31 ng/mg), lidocaine (34 pg/mg), diazepam (3 pg/mg) and its metabolite nordazepam (5 pg/mg), and codeine (16 pg/mg) in the analyzed hair of child G. The presence of diazepam and its metabolite can be explained by medical care. The low concentration of codeine without its metabolite morphine, may suggest an external contamination by the environment, contact with people who had consumed this molecule (bedding, caressing, etc.), especially the child's father. The presence of metoclopramide in the blood and hair is compatible with a passage of breast milk to the child. According to the literature, less than 5% of the maternal dose is found in the child. Adverse effects such as intestinal disorders, drowsiness, agitation, hypertonia have already been described in breastfed children (EMA. Metoclopramide Assessment report, 2013). This case illustrates the passage into breast milk of rectally administered metoclopramide in the mother and the toxic effects of this molecule in the child. The interpretation of hair analysis in infants remains nevertheless delicate to discriminate consumption from environmental contamination.

Pharmacokinetic Drug Interaction Study of Sorafenib and Morphine in Rats.

A combination of the tyrosine kinase inhibitor—sorafenib—and the opioid analgesic—morphine—can be found in the treatment of cancer patients. Since both are substrates of P-glycoprotein (P-gp), and sorafenib is also an inhibitor of P-gp, their co-administration may affect their pharmacokinetics, and thus the safety and efficacy of cancer therapy. Therefore, the aim of this study was to evaluate the potential pharmacokinetic drug–drug interactions between sorafenib and morphine using an animal model. The rats were divided into three groups that Received: sorafenib and morphine (ISOR+MF), sorafenib (IISOR), and morphine (IIIMF). Morphine caused a significant increase in maximum plasma concentrations (Cmax) and the area under the plasma concentration–time curves (AUC0–t, and AUC0–∞) of sorafenib by 108.3 (p = 0.003), 55.9 (p = 0.0115), and 62.7% (p = 0.0115), respectively. Also, the Cmax and AUC0–t of its active metabolite—sorafenib N-oxide—was significantly increased in the presence of morphine (p = 0.0022 and p = 0.0268, respectively). Sorafenib, in turn, caused a significant increase in the Cmax of morphine (by 0.5-fold, p = 0.0018). Moreover, in the presence of sorafenib the Cmax, AUC0–t, and AUC0–∞ of the morphine metabolite M3G increased by 112.62 (p < 0.0001), 46.82 (p = 0.0124), and 46.78% (p = 0.0121), respectively. Observed changes in sorafenib and morphine may be of clinical significance. The increased exposure to both drugs may improve the response to therapy in cancer patients, but on the other hand, increase the risk of adverse effects.

Should Codeine Still be Considered a WHO Essential Medicine?

Codeine continues to be widely used as an analgesic, antidiarrhoeal and antitussive agent. Its analgesic effect depends on its biotransformation to morphine, a strong opioid. The highly variable biotransformation of codeine to morphine, catalysed by CYP2D6, underlies the pronounced interindividual variability of its analgesic response. Randomized controlled trials have demonstrated that codeine administered alone has the poorest analgesic effect among all commonly used analgesics in acute postoperative pain. Moreover, it is highly unlikely that the low dose of codeine contributes to the pain-relieving effect of the non-opioid component in combination analgesic products. In addition, there is a lack of reliable clinical evidence to support the use of codeine as an antitussive in acute or chronic cough. Codeine use, through its active metabolite morphine, has the potential to lead to abuse and dependence. The World Health Organization (WHO) removed codeine from the essential medicines list for children in 2011. Based on the available information in the scientific literature on the efficacy and safety of codeine, the WHO should seriously consider removing it also from the list of essential medicines for adults, which would be a strong signal for all health professionals to prescribe and dispense codeine with the utmost caution.

Functional phenotyping of the CYP2D6 probe drug codeine in the horse

BackgroundIn humans, the drug metabolizing enzyme CYP2D6 is highly polymorphic resulting in substantial differences in the metabolism of drugs including anti-arrhythmics, neuroleptics, and opioids. The objective of this study was to phenotype a population of 100 horses from five different breeds and assess differences in the metabolic activity of the equine CYP2D6 homolog using codeine as a probe drug. Administration of a probe drug is a common method used for patient phenotyping in human medicine, whereby the ratio of parent drug to metabolite (metabolic ratio, MR) can be used to compare relative enzyme function between individuals. A single oral dose of codeine (0.6 mg/kg) was administered and plasma concentrations of codeine and its metabolites were determined using liquid chromatography mass spectrometry. The MR of codeine O-demethylation [(codeine)/(morphine + morphine-3-glucuronide + morphine-6-glucuronide)] was determined using the area under the plasma concentration-time curve extrapolated from time zero to infinity (AUC0-∞) for each analyte and used to group horses into predicted phenotypes (high-, moderate-, and low-MR).ResultsThe MR of codeine O-demethylation ranged from 0.002 to 0.147 (median 0.018) among all horses. No significant difference in MR was observed between breeds, age, or sex. Of the 100 horses, 11 were classified as high-MR, 72 moderate-MR, and 17 low-MR. Codeine AUC0-∞ and O-demethylation MR were significantly different (p < 0.05) between all three groups. The mean ± SD MR was 0.089 ± 0.027, 0.022 ± 0.011, and 0.0095 ± 0.001 for high-, moderate-, and low-MR groups, respectively. The AUC for the morphine metabolites morphine-3-glucuronide and morphine-6-glucuronide were significantly different between high-and low-MR groups (p < 0.004 and p < 0.006).ConclusionsThe MR calculated from plasma following codeine administration allowed for classification of horses into metabolic phenotypes within a large population. The range of codeine metabolism observed among horses suggests the presence of genetic polymorphisms in CYP2D82 of which codeine is a known substrate. Additional studies including CYP2D82 genotyping of high- and low-MR individuals are necessary to determine the presence of CYP2D polymorphisms and their functional implications with respect to the metabolism of therapeutics.

Systematic Structure-Based Virtual Screening Approach to Antibody Selection and Design of a Humanized Antibody against Multiple Addictive Opioids without Affecting Treatment Agents Naloxone and Naltrexone

Opioid drug use, especially heroin, is known as a growing national crisis in America. Heroin itself is a prodrug and is converted to the most active metabolite 6-monoacetylmorphine (6-MAM) responsible for the acute toxicity of heroin and then to a relatively less-active metabolite morphine responsible for the long-term toxicity of heroin. Monoclonal antibodies (mAbs) are recognized as a potentially promising therapeutic approach in the treatment of opioid use disorders (OUDs). Due to the intrinsic challenges of discovering an mAb against multiple ligands, here we describe a general, systematic structure-based virtual screening and design approach which has been used to identify a known anti-morphine antibody 9B1 and a humanized antibody h9B1 capable of binding to multiple addictive opioids (including 6-MAM, morphine, heroin, and hydrocodone) without significant binding with currently available OUD treatment agents naloxone, naltrexone, and buprenorphine. The humanized antibody may serve as a promising candidate for the treatment of OUDs. The experimental binding affinities reasonably correlate with the computationally predicted binding free energies. The experimental activity data strongly support the computational predictions, suggesting that the systematic structure-based virtual screening and humanization design protocol is reliable. The general, systematic structure-based virtual screening and design approach will be useful for many other antibody selection and design efforts in the future.

Morphine‐6‐Glucuronide Isomers‐Synthesis and Biological Evaluation

Morphine‐6β‐D‐glucuronide (M6βG), an active metabolite of morphine, and its isomer morphine‐6α‐D‐glucuronide (M6αG) were synthesized from 3‐O‐protected morphinethrough glycosylation and alkaline hydrolysis. All structures were determined by spectroscopic analysis, and especially it is the first time to report the single crystal of compound M6αG. In vitro binding assay showed that M6βG bound to mu opioid receptor (MOR), kappa opioid receptor (KOR), and delta opioid receptor (DOR) with nanomolar affinity (Ki = 28.03, 116.88, and 375.13 nM) and M6αG bound to them with similar affinity (Ki = 1070.13, 20 637.93, and 677.36 nM). The selectivity of M6αG toward KOR is much higher. Hot‐plate test showed that the analgesic effect of M6βG is better than that of M6αG, that is maybe because the mechanism of M6αG is different.

Glutathione and Glutathione-Like Sequences of Opioid and Aminergic Receptors Bind Ascorbic Acid, Adrenergic and Opioid Drugs Mediating Antioxidant Function: Relevance for Anesthesia and Abuse.

Opioids and their antagonists alter vitamin C metabolism. Morphine binds to glutathione (l-γ-glutamyl-l-cysteinyl-glycine), an intracellular ascorbic acid recycling molecule with a wide range of additional activities. The morphine metabolite morphinone reacts with glutathione to form a covalent adduct that is then excreted in urine. Morphine also binds to adrenergic and histaminergic receptors in their extracellular loop regions, enhancing aminergic agonist activity. The first and second extracellular loops of adrenergic and histaminergic receptors are, like glutathione, characterized by the presence of cysteines and/or methionines, and recycle ascorbic acid with similar efficiency. Conversely, adrenergic drugs bind to extracellular loops of opioid receptors, enhancing their activity. These observations suggest functional interactions among opioids and amines, their receptors, and glutathione. We therefore explored the relative binding affinities of ascorbic acid, dehydroascorbic acid, opioid and adrenergic compounds, as well as various control compounds, to glutathione and glutathione-like peptides derived from the extracellular loop regions of the human beta 2-adrenergic, dopamine D1, histamine H1, and mu opioid receptors, as well as controls. Some cysteine-containing peptides derived from these receptors do bind ascorbic acid and/or dehydroascorbic acid and the same peptides generally bind opioid compounds. Glutathione binds not only morphine but also naloxone, methadone, and methionine enkephalin. Some adrenergic drugs also bind to glutathione and glutathione-like receptor regions. These sets of interactions provide a novel basis for understanding some ways that adrenergic, opioid and antioxidant systems interact during anesthesia and drug abuse and may have utility for understanding drug interactions.

Fentanyl but not Morphine Interacts with Nonopioid Recombinant Human Neurotransmitter Receptors and Transporters.

Synthetic opioids, including fentanyl and its analogs, have therapeutic efficacy in analgesia and anesthesia. However, their illicit use in the United States has increased and contributed to the number one cause of death for adults 18-50 years old. Fentanyl and the heroin metabolite morphine induce respiratory depression that can be treated with the μ opioid receptor (MOR) antagonist naloxone. With higher or more rapid dosing, fentanyl, more than morphine, causes chest wall rigidity and can also induce rapid onset laryngospasm. Because non-MORs could mediate differing clinical manifestations, we examined the interactions of fentanyl and morphine at recombinant human neurotransmitter transporters, G protein-coupled receptors, and the N-methyl-D-aspartate glutamate receptor. Both drugs were agonists at MOR, κ, and δ opioid receptors. Morphine had little or no affinity at other human receptors and transporters (K i or IC50 value >100 µM). However, fentanyl had K i values of 1407 and 1100 nM at α 1A and α 1B adrenoceptor subtypes, respectively, and K i values of 1049 and 1670 nM at dopamine D4.4 and D1 receptor subtypes, respectively; it also blocked [3H]neurotransmitter uptake by the vesicular monoamine transporter 2 (IC50 = 911 nM). Pharmacokinetic models indicate that these Ki and IC50 values are pharmacologically relevant. Fentanyl had little affinity for other receptors or transporters. Thus, noradrenergic disposition at specific receptor subtypes in relevant organs may play a role in respiratory and cardiothoracic effects of fentanyl. Data suggest that less selective fentanyl receptor pharmacology could play a role in the different clinical effects of morphine compared with fentanyl, including fentanyl-induced deaths after illicit use. SIGNIFICANCE STATEMENT: The synthetic opioid fentanyl induces different clinical effects, including rapid onset muscular rigidity, vocal cord closure, and rapid death, than the heroin metabolite morphine. Our data indicate for the first time that the two drugs have very different effects at recombinant human neurotransmitter receptors and transporters that might explain those clinical differences.

Metabolism, pharmacokinetics and selected pharmacodynamic effects of codeine following a single oral administration to horses

ObjectiveTo describe the pharmacokinetics and selected pharmacodynamic variables of codeine and its metabolites in Thoroughbred horses following a single oral administration. Study designProspective experimental study. AnimalsA total of 12 Thoroughbred horses, nine geldings and three mares, aged 4–8 years. MethodsHorses were administered codeine (0.6 mg kg–1) orally and blood was collected before administration and at various times until 120 hours post administration. Plasma and urine samples were collected and analyzed for codeine and its metabolites by liquid chromatography–mass spectrometry, and plasma pharmacokinetics were determined. Heart rate and rhythm, step counts, packed cell volume and total plasma protein were measured before and 4 hours after administration. ResultsCodeine was rapidly converted to the metabolites norcodeine, codeine-6-glucuronide (C6G), morphine, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G). Plasma codeine concentrations were best represented using a two-compartment model. The Cmax, tmax and elimination t½ were 270.7 ± 136.0 ng mL–1, 0.438 ± 0.156 hours and 2.00 ± 0.534 hours, respectively. M3G was the main metabolite detected (Cmax 492.7 ± 35.5 ng mL–1), followed by C6G (Cmax 96.1 ± 33.8 ng mL–1) and M6G (Cmax 22.3 ± 4.96 ng mL–1). Morphine and norcodeine were the least abundant metabolites with Cmax of 3.17 ± 0.95 and 1.42 ± 0.79 ng mL–1, respectively. No significant adverse or excitatory effects were observed. Conclusions and clinical relevanceFollowing oral administration, codeine is rapidly metabolized to morphine, M3G, M6G, C6G and norcodeine in horses. Plasma concentrations of M6G, a presumed active metabolite of morphine, were comparable to concentrations reported previously following administration of an analgesic dose of morphine to horses. Codeine was well tolerated based on pharmacodynamic variables and behavioral observations.

Morphine-3-glucuronide causes antinociceptive cross-tolerance to morphine and increases spinal substance P expression

Morphine-3-glucuronide (M3G), the main metabolite of morphine, has been implicated in the development of tolerance and of opioid-induced hyperalgesia, both limiting the analgesic use of morphine. We evaluated the acute and chronic effects of M3G and morphine as well as development of antinociceptive cross-tolerance between morphine and M3G after intrathecal administration and assessed the expression of pain-associated neurotransmitter substance P in the spinal cord. Sprague-Dawley rats received intrathecal M3G or morphine twice daily for 6 days. Nociception and tactile allodynia were measured with von Frey filaments after acute and chronic treatments. Substance P levels in the dorsal horn of the spinal cord were determined by immunohistochemistry after 4-day treatments. Acute morphine caused antinociception as expected, whereas acute M3G caused tactile allodynia, as did both chronic M3G and morphine. Chronic M3G also induced antinociceptive cross-tolerance to morphine. M3G and morphine increased substance P levels similarly in the nociceptive laminae of the spinal cord. This study shows that chronic intrathecal M3G sensitises animals to mechanical stimulation and elevates substance P levels in the nociceptive laminae of the spinal cord. Chronic M3G also induces antinociceptive cross-tolerance to morphine. Thus, chronic M3G exposure might contribute to morphine-induced tolerance and opioid-induced hyperalgesia.

Hetero-oligomer formation of mouse UDP-glucuronosyltransferase (UGT) 2b1 and 1a1 results in the gain of glucuronidation activity towards morphine, an activity which is absent in homo-oligomers of either UGT

UDP-Glucuronosyltransferase (UGT, Ugt) is a major drug metabolizing enzyme family involved in the glucuronidation and subsequent elimination of drugs and small lipophilic molecules. UGT forms homo- and hetero-oligomers that enhance or suppress UGT activity. In our previous study, we characterized mouse Ugt1a1 and all the Ugt isoform belonging to the Ugt2b subfamily and revealed that mouse Ugt2b1 and Ugt1a1 cannot metabolize morphine. Mouse Ugt2b1 had been believed to function similarly to rat UGT2B1, which plays a major role in morphine glucuronidation in rat liver. Thus, in this study, we hypothesized that hetero-oligomerization with another Ugt isoform may affect Ugt2b1 catalytic ability. We co-expressed Ugt1a1 and Ugt2b1 in a baculovirus-insect cell system, and confirmed hetero-oligomer formation by co-immunoprecipitation. As reported previously, microsomes singly expressing Ugt1a1 or Ugt2b1 were inactive towards the glucuronidation of morphine. Interestingly, in contrast, morphine-3-glucuronide, a major metabolite of morphine was formed, when Ugt2b1 and Ugt1a1 were co-expressed. This effect of hetero-oligomerization of Ugt1a1 and Ugt2b1 was also observed for 17β-estradiol glucuronidation. This is the first report demonstrating that UGT acquires a novel catalytic ability by forming oligomers. Protein-protein interaction of Ugts may contribute to robust detoxification of xenobiotics by altering the substrate diversity of the enzymes.