Mu-opioid Research Articles

βγ G-proteins, but not regulators of G-protein signaling 4, modulate opioid-induced respiratory rate depression.

Opioid medications are the mainstay of pain management but present substantial side-effects such as respiratory depression which can be lethal with overdose. Most opioid drugs, such as fentanyl, act on opioid receptors such as the G-protein-coupled µ-opioid receptors (MOR). G-protein-coupled receptors activate pertussis toxin-sensitive G-proteins to inhibit neuronal activity. Binding of opioid ligands to MOR and subsequent activation G proteins βγ is modulated by regulator of G-protein signaling (RGS). The roles of G-proteins βγ and RGS in MOR-mediated inhibition of the respiratory network are not known. Using rodent models to pharmacologically modulate G-protein signaling, we aim to determine the roles of βγ G-proteins and RGS4. We showed that inhibition of βγ G-proteins using gallein perfused in the brainstem circuits regulating respiratory depression by opioid drugs results in complete reversal of respiratory depression. Blocking of RGS4 using CCG55014 did not change the respiratory depression induced by MOR activation despite co-expression of RGS4 and MORs in the brainstem. Our results suggest that neuronal inhibition by opioid drugs is mediated by G-proteins, but not by RGS4, which supports the concept that βγ G-proteins could be molecular targets to develop opioid overdose antidotes without the risks of re-narcotization often found with highly potent opioid drugs. On the other hand, RGS4 mediates opioid analgesia, but not respiratory depression, and RGS4 may be molecular targets to develop pain therapies without respiratory liability.

Pharmacokinetic and Pharmacodynamic Consequences of Cytochrome P450 3A Inhibition on Mitragynine Metabolism in Rats.

Mitragynine, an opioidergic alkaloid present in Mitragyna speciosa (kratom), is metabolized by cytochrome P450 3A (CYP3A) to 7-hydroxymitragynine, a more potent opioid receptor agonist. The extent to which conversion to 7-hydroxymitragynine mediates the in vivo effects of mitragynine is unclear. The current study examined how CYP3A inhibition (ketoconazole) modifies the pharmacokinetics of mitragynine in rat liver microsomes in vitro. The study further examined how ketoconazole modifies the discriminative stimulus and antinociceptive effects of mitragynine in rats. Ketoconazole [30 mg/kg, oral gavage (o.g.)] increased systemic exposure to mitragynine (13.3 mg/kg, o.g.) by 120% and 7-hydroxymitragynine exposure by 130%. The unexpected increase in exposure to 7-hydroxymitragynine suggested that ketoconazole inhibits metabolism of both mitragynine and 7-hydroxymitragynine, a finding confirmed in rat liver microsomes. In rats discriminating 3.2 mg/kg morphine from vehicle under a fixed-ratio schedule of food delivery, ketoconazole pretreatment increased the potency of both mitragynine (4.7-fold) and 7-hydroxymitragynine (9.7-fold). Ketoconazole did not affect morphine's potency. Ketoconazole increased the antinociceptive potency of 7-hydroxymitragynine by 4.1-fold. Mitragynine (up to 56 mg/kg, i.p.) lacked antinociceptive effects both in the presence and absence of ketoconazole. These results suggest that both mitragynine and 7-hydroxymitragynine are cleared via CYP3A and that 7-hydroxymitragynine is formed as a metabolite of mitragynine by other routes. These results have implications for kratom use in combination with numerous medications and citrus juices that inhibit CYP3A. SIGNIFICANCE STATEMENT: Mitragynine is an abundant kratom alkaloid that exhibits low efficacy at the μ-opioid receptor (MOR). Its metabolite, 7-hydroxymitragynine, is also an MOR agonist but with higher affinity and efficacy than mitragynine. Our results in rats demonstrate that cytochrome P450 3A (CYP3A) inhibition can increase the systematic exposure of both mitragynine and 7-hydroxymitragynine and their potency to produce MOR-mediated behavioral effects. These data highlight potential interactions between kratom and CYP3A inhibitors, which include numerous medications and citrus juices.

Engineering human spinal microphysiological systems to model opioid-induced tolerance.

pioids are commonly used for treating chronic pain. However, with continued use, they may induce tolerance and/or hyperalgesia, which limits therapeutic efficacy. The human mechanisms of opioid-induced tolerance and hyperalgesia are significantly understudied, in part, because current models cannot fully recapitulate human pathology. Here, we engineered novel human spinal microphysiological systems (MPSs) integrated with plug-and-play neural activity sensing for modeling human nociception and opioid-induced tolerance. Each spinal MPS consists of a flattened human spinal cord organoid derived from human stem cells and a 3D printed organoid holder device for plug-and-play neural activity measurement. We found that the flattened organoid design of MPSs not only reduces hypoxia and necrosis in the organoids, but also promotes their neuron maturation, neural activity, and functional development. We further demonstrated that prolonged opioid exposure resulted in neurochemical correlates of opioid tolerance and hyperalgesia, as measured by altered neural activity, and downregulation of μ-opioid receptor expression of human spinal MPSs. The MPSs are scalable, cost-effective, easy-to-use, and compatible with commonly-used well-plates, thus allowing plug-and-play measurements of neural activity. We believe the MPSs hold a promising translational potential for studying human pain etiology, screening new treatments, and validating novel therapeutics for human pain medicine.

The effect of dalargin on growth factors content in experimental ulcerative colitis

Introduction: The effectiveness of ulcerative colitis (UC) treatment is fairy moderate and it gives a rise to search for new ways of treating it. Considering the combination of dalargineffects, studying the dalargin influence on the UC development is of undoubted interest. The aim of the study was to evaluate the dalargin effect on the content of transforming growth factor-β (TGF-β) and epidermal growth factor (EGF) in the colonic wall in mice with experimental ulcerative colitis. Materials and methods: UC was simulated by replacing water with a 5% solution of dextran sodium sulfate in boiled water for 5 days. The mice were killed on the 5th, 7th and 28th days. The concentrations of TGF-β and EGF in the homogenate of the medial colon were determined by enzyme-linked immunosorbent assay using standard kits. Results and discussion: The dalargin daily subcutaneous administration (dose of 100 μg/kg) for 7 days led to a decrease in TGF-β levels on the 5th and 7th days compared to the control group. In chronic UC, the concentration of TGF-β was higher than in the control group. The EGF concentration was increased in mice with UC treated with dalargin throughout the experiment. There were no differences in dalargin and sulfasalazine effects on the content of TGF-β, and the concentration of EGF throughout the experiment was significantly higher in the animals treated with dalargin. Conclusion: Effect of dalargin on the TGF-β and EGF concentrations was explained by its stimulating action to opioid μ-receptors localized on immune cells of the colon.

Structure-Kinetics Relationships of Opioids from Metadynamics and Machine Learning Analysis.

The nation's opioid overdose deaths reached an all-time high in 2021. The majority of deaths are due to synthetic opioids represented by fentanyl. Naloxone, which is a FDA-approved reversal agent, antagonizes opioids through competitive binding at the μ-opioid receptor (mOR). Thus, knowledge of the opioid's residence time is important for assessing the effectiveness of naloxone. Here, we estimated the residence times (τ) of 15 fentanyl and 4 morphine analogs using metadynamics and compared them with the most recent measurement of the opioid kinetic, dissociation, and naloxone inhibitory constants (Mann et al. Clin. Pharmacol. Therapeut. 2022, 120, 1020-1232). Importantly, the microscopic simulations offered a glimpse at the common binding mechanism and molecular determinants of dissociation kinetics for fentanyl analogs. The insights inspired us to develop a machine learning approach to analyze the kinetic impact of fentanyl's substituents based on the interactions with mOR residues. This proof-of-concept approach is general; for example, it may be used to tune ligand residence times in computer-aided drug discovery.

Analgesic Effect of Buprenorphine for Chronic Noncancer Pain: A Systematic Review and Meta-analysis of Randomized Controlled Trials.

Buprenorphine is a partial agonist at the µ-opioid receptor and an antagonist at the delta and kappa opioid receptors. It has high affinity and low intrinsic activity at the µ-opioid receptor. Buprenorphine demonstrates no ceiling effect for clinical analgesia, but demonstrates this for respiratory depression and euphoria. It may provide effective analgesia while producing less adverse effects, making it a promising opioid analgesic. A systematic review and meta-analysis were performed to examine the analgesic efficacy of buprenorphine for patients with chronic noncancer pain. PubMed, MEDLNE, Embase, and the Cochrane Library were searched up to January 2022. Randomized controlled trials were included if they compared buprenorphine versus placebo or active analgesic in patients with chronic noncancer pain, where pain score was an outcome. Nonrandomized controlled trials, observational studies, qualitative studies, case reports, and commentaries were excluded. Two investigators independently performed the literature search, study selection, and data collection. A random-effects model was used. The primary outcome was the effect of buprenorphine on pain intensity in patients with chronic noncancer pain based on standardized mean difference (SMD) in pain score. Quality of evidence was assessed using the Grade of Recommendations Assessment, Development, and Evaluation (GRADE) approach. Two separate literature searches were conducted for patients with and without opioid use disorder (OUD). Only one study met the search criteria for those with OUD. Fourteen randomized controlled trials were included for those without OUD. Buprenorphine was associated with reduced pain score (SMD = -0.368, P < .001, I2 = 89.37%) compared to placebo or active analgesic. Subgroup meta-analyses showed statistically significant differences in favor of buprenorphine versus placebo (SMD = -0.404, P < .001), for chronic low back pain (SMD = -0.383, P < .001), when administered via the transdermal route (SMD = -0.572, P = .001), via the buccal route (SMD = -0.453, P < .001), with length of follow-up lasting <12 weeks (SMD = -0.848, P < .05), and length of follow-up lasting 12 weeks or more (SMD = -0.415, P < .001). There was no significant difference when compared to active analgesic (SMD = 0.045, P > .05). Quality of evidence was low to moderate. Buprenorphine was associated with a statistically significant and small reduction in pain intensity compared to placebo. Both the transdermal and buccal routes provided pain relief. There was more evidence supporting its use for chronic low back pain.

The safety and efficacy of methylnaltrexone in pediatric oncology patients: A single center experience.

Peripherally acting μ-opioid receptor antagonists (PAMORAs) are used in the treatment of opioid induced constipation without impacting the actions of opioid analgesics. Subcutaneous methylnaltrexone was one of the first PAMORAs approved in April 2008 for the treatment of opioid induced constipation in adult patients. The safety and effectiveness of methylnaltrexone has not been established in pediatric patients. In this study, the use of subcutaneous methylnaltrexone in pediatric patients is analyzed and reviewed. The primary outcome is occurrence of a bowel movement within 24 h after methylnaltrexone (MNTX) administration and the number of bowel movements following treatment with methylnaltrexone. Secondary outcomes include safety in this patient cohort. This is a retrospective study of 79 pediatric patients with opioid induced constipation. Patients were administered methylnaltrexone during their inpatient stay. Data on bowel activity after methylnaltrexone was obtained from the hospital information system. Out of the 79 patients who received methylnaltrexone, there were seven patients from whom data could not be analyzed. Of the 72 patients whose data was available, 38% (N = 27) were documented as having a bowel movement, 62% (N = 45) did not have a bowel movement. Reported adverse events were minimal with nausea (N = 3), vomiting (N = 1), and flatulence (N = 6). Methylnaltrexone appears safe in the pediatric population and produces bowel movements in more than a third of pediatric patients. It is a feasible and safe option for opioid induced constipation in pediatric patients.

Combined non-psychoactive Cannabis components cannabidiol and β-caryophyllene reduce chronic pain via CB1 interaction in a rat spinal cord injury model.

The most frequently reported use of medical marijuana is for pain relief. However, its psychoactive component Δ9-tetrahydrocannabinol (THC) causes significant side effects. Cannabidiol (CBD) and β-caryophyllene (BCP), two other cannabis constituents, possess more benign side effect profiles and are also reported to reduce neuropathic and inflammatory pain. We evaluated the analgesic potential of CBD and BCP individually and in combination in a rat spinal cord injury (SCI) clip compression chronic pain model. Individually, both phytocannabinoids produced dose-dependent reduction in tactile and cold hypersensitivity in male and female rats with SCI. When co-administered at fixed ratios based on individual A50s, CBD and BCP produced enhanced dose-dependent reduction in allodynic responses with synergistic effects observed for cold hypersensitivity in both sexes and additive effects for tactile hypersensitivity in males. Antinociceptive effects of both individual and combined treatment were generally less robust in females than males. CBD:BCP co-administration also partially reduced morphine-seeking behavior in a conditioned place preference (CPP) test. Minimal cannabinoidergic side effects were observed with high doses of the combination. The antinociceptive effects of the CBD:BCP co-administration were not altered by either CB2 or μ-opioid receptor antagonist pretreatment but, were nearly completely blocked by CB1 antagonist AM251. Since neither CBD or BCP are thought to mediate antinociception via CB1 activity, these findings suggest a novel CB1 interactive mechanism between these two phytocannabinoids in the SCI pain state. Together, these findings suggest that CBD:BCP co-administration may provide a safe and effective treatment option for the management of chronic SCI pain.

A277 A NOVEL PH-SENSITIVE Μ-OPIOID RECEPTOR AGONIST THAT DOES NOT INDUCE TOLERANCE IN A COLITIS MOUSE MODEL

Abstract Background Adequate pain control in inflammatory bowel disease (IBD) can require opioids due to their high analgesic potency. The long-term use of opioids, however, is limited by the development of tolerance. This leads to reduced analgesic efficacy over time, resulting in escalating opioid dosing and thus increased risk of serious side effects. We previously demonstrated the safety and effectiveness of a novel pH-sensitive µ-opioid receptor (MOR) agonist, NFEPP, in a murine colitis pain model, but its tolerance potential with chronic administration is unknown. Purpose To assess the tolerance potential of NFEPP compared to its parent compound fentanyl during acute colitis in a preclinical mouse model. Method Acute colitis in C57BL/6 mice was induced using 2.5% dextran sulphate sodium for 5 days. NFEPP or fentanyl were then administered s.c. every 4 hours between 7am and 11pm over 5 days in daily increasing concentrations (0.4-1.5 mg/kg/d). Analgesic tolerance to opioids was assessed in conscious mice by measuring visceromotor responses (VMRs) to noxious colorectal distensions. Tolerance to the MOR agonist DAMGO in NFEPP or fentanyl treated mice was evaluated using patch-clamp recordings from dorsal root ganglion (DRG) neurons and extracellular recordings from lumbar splanchnic nerves that innervate the colon. Inflammation was assessed by macroscopic analyses, histological scoring and tissue-pH measurements of the inflamed colon. Group differences were analyzed using two-way ANOVA with Bonferroni′s or Tukey′s post-test (p-value &amp;lt;0.05). Result(s) NFEPP significantly reduced VMRs before and after chronic NFEPP treatment (39% reduction, p&amp;lt;0.05 vs. 41% reduction, p&amp;lt;0.05, compared to baseline at 80 µl). No differences of NFEPP induced antinociceptive actions were observed comparing VMR measurements before and after chronic administration (p=0.44). However, the analgesic activity of fentanyl decreased over time with less VMR inhibition observed after chronic treatment compared to fentanyl naïve mice (p&amp;lt;0.05). Cross tolerance testing between these two opioids revealed a loss of NFEPP- induced VMR inhibition in fentanyl treated mice, but fentanyl effects in the NFEPP treatment group remained unchanged. In patch-clamp recordings, DAMGO (100nM) evoked antinociceptive actions in DRG neurons from NFEPP treated mice (rheobase increase: 50%, p&amp;lt;0.05, compared to vehicle) whereas no change in rheobase with DAMGO was observed in DRG neurons from fentanyl treated mice. Afferent nerve activity from lumbar splanchnic nerves in response to von Frey filament probing decreased after DAMGO (100nM) application in NFEPP treated mice (p&amp;lt;0.05, compared to control probing), but showed no change in fentanyl treated mice. Colonic length, diameter, histological damage and tissue-pH did not differ between the two opioid treatment groups. Conclusion(s) Chronic administration of NFEPP during acute murine colitis does not lead to analgesic tolerance within the visceral nociceptive system, in contrast to its parent compound fentanyl. Please acknowledge all funding agencies by checking the applicable boxes below CCC Disclosure of Interest None Declared

Application of Mixed-Solvent Molecular Dynamics Simulations for Prediction of Allosteric Sites on G Protein-Coupled Receptors.

The development of small molecule allosteric modulators acting at G protein-coupled receptors (GPCRs) is becoming increasingly attractive. Such compounds have advantages over traditional drugs acting at orthosteric sites on these receptors, in particular target specificity. However, the number and locations of druggable allosteric sites within most clinically relevant GPCRs are unknown. In the present study, we describe the development and application of a mixed-solvent molecular dynamics (MixMD)-based method for the identification of allosteric sites on GPCRs. The method employs small organic probes with druglike qualities to identify druggable hotspots in multiple replicate short-timescale simulations. As proof of principle, we first applied the method retrospectively to a test set of five GPCRs (cannabinoid receptor type 1, C-C chemokine receptor type 2, M2 muscarinic receptor, P2Y purinoceptor 1, and protease-activated receptor 2) with known allosteric sites in diverse locations. This resulted in the identification of the known allosteric sites on these receptors. We then applied the method to the μ-opioid receptor. Several allosteric modulators for this receptor are known, although the binding sites for these modulators are not known. The MixMD-based method revealed several potential allosteric sites on the mu-opioid receptor. Implementation of the MixMD-based method should aid future efforts in the structure-based drug design of drugs targeting allosteric sites on GPCRs. SIGNIFICANCE STATEMENT: Allosteric modulation of G protein-coupled receptors (GPCRs) has the potential to provide more selective drugs. However, there are limited structures of GPCRs bound to allosteric modulators, and obtaining such structures is problematic. Current computational methods utilize static structures and therefore may not identify hidden or cryptic sites. Here we describe the use of small organic probes and molecular dynamics to identify druggable allosteric hotspots on GPCRs. The results reinforce the importance of protein dynamics in allosteric site identification.

Phosphorylated Upstream Frameshift 1-dependent Nonsense-mediated μ-Opioid Receptor mRNA Decay in the Spinal Cord Contributes to the Development of Neuropathic Allodynia-like Behavior in Rats.

Nonsense-mediated messenger RNA (mRNA) decay increases targeted mRNA degradation and has been implicated in the regulation of gene expression in neurons. The authors hypothesized that nonsense-mediated μ-opioid receptor mRNA decay in the spinal cord is involved in the development of neuropathic allodynia-like behavior in rats. Adult Sprague-Dawley rats of both sexes received spinal nerve ligation to induce neuropathic allodynia-like behavior. The mRNA and protein expression contents in the dorsal horn of animals were measured by biochemical analyses. Nociceptive behaviors were evaluated by the von Frey test and the burrow test. On Day 7, spinal nerve ligation significantly increased phosphorylated upstream frameshift 1 (UPF1) expression in the dorsal horn (mean ± SD; 0.34 ± 0.19 in the sham ipsilateral group vs. 0.88 ± 0.15 in the nerve ligation ipsilateral group; P < 0.001; data in arbitrary units) and drove allodynia-like behaviors in rats (10.58 ± 1.72 g in the sham ipsilateral group vs. 1.19 ± 0.31 g in the nerve ligation ipsilateral group, P < 0.001). No sex-based differences were found in either Western blotting or behavior tests in rats. Eukaryotic translation initiation factor 4A3 (eIF4A3) triggered SMG1 kinase (0.06 ± 0.02 in the sham group vs. 0.20 ± 0.08 in the nerve ligation group, P = 0.005, data in arbitrary units)-mediated UPF1 phosphorylation, leading to increased nonsense-mediated mRNA decay factor SMG7 binding and µ-opioid receptor mRNA degradation (0.87 ± 0.11-fold in the sham group vs. 0.50 ± 0.11-fold in the nerve ligation group, P = 0.002) in the dorsal horn of the spinal cord after spinal nerve ligation. Pharmacologic or genetic inhibition of this signaling pathway in vivo ameliorated allodynia-like behaviors after spinal nerve ligation. This study suggests that phosphorylated UPF1-dependent nonsense-mediated μ-opioid receptor mRNA decay is involved in the pathogenesis of neuropathic pain.

Synthesis, structure, in vitro pharmacology, and in vivo activity of trans-3,4-dichloro-N-[[1-(dimethylamino)-4-phenylcyclohexyl]methyl]-benzamide (AP01; 4-phenyl-AH-7921), a novel mixed μ-/κ-opioid.

Analogs of non-fentanyl novel synthetic opioids (NSO) with modifications that fall outside of established structure-activity relationships (SARs) for that class of drugs create the question whether or not it should be considered an analog, as defined by 21 U.S.C. §802(32)(A), which is important for its inclusion in the US system of drug scheduling. AH-7921 is a US Schedule I drug and an example of the 1-benzamidomethyl-1-cyclohexyldialkylamine class of NSO. The SARs regarding substitution of the central cyclohexyl ring have not been well characterized in the literature. Therefore, in order to expand the SAR surrounding AH-7921 analogs, trans-3,4-dichloro-N-[[1-(dimethylamino)-4-phenylcyclohexyl]methyl]-benzamide (AP01; 4-phenyl-AH-7921) has been synthesized, analytically characterized, and tested in vitro and in vivo pharmacologically. Using methods described in the original patents for this class of NSO, it was found that the single trans geometric isomer was obtained. The proton nuclear magnetic resonance, mass spectrum, infrared spectrum, and Raman spectrum are reported along with the melting point of the hydrochloride salt. In vitro binding to a battery of 43 central nervous system receptors showed it to be a high-affinity μ-opioid receptor (MOR) and κ-opioid receptor (KOR) ligand (60 nM and 34 nM, respectively). AP01 also had a 4nM affinity for the serotonin transporter (SERT), which is a higher level of potency at this receptor than most other opioids. In rats, it exhibited antinociception in the acetic acid writhing test. Therefore, the 4-phenyl modification results in an active NSO, but carries with it potential toxicities beyond those expected for currently approved opioid drugs.

All-Atom Molecular Dynamics Simulations Indicated the Involvement of a Conserved Polar Signaling Channel in the Activation Mechanism of the Type I Cannabinoid Receptor.

The type I cannabinoid G protein-coupled receptor (CB1, GPCR) is an intensely investigated pharmacological target, owing to its involvement in numerous physiological functions as well as pathological processes such as cancers, neurodegenerative diseases, metabolic disorders and neuropathic pain. In order to develop modern medications that exert their effects through binding to the CB1 receptor, it is essential to understand the structural mechanism of activation of this protein. The pool of atomic resolution experimental structures of GPCRs has been expanding rapidly in the past decade, providing invaluable information about the function of these receptors. According to the current state of the art, the activity of GPCRs involves structurally distinct, dynamically interconverting functional states and the activation is controlled by a cascade of interconnecting conformational switches in the transmembrane domain. A current challenge is to uncover how different functional states are activated and what specific ligand properties are responsible for the selectivity towards those different functional states. Our recent studies of the μ-opioid and β2-adrenergic receptors (MOP and β2AR, respectively) revealed that the orthosteric binding pockets and the intracellular surfaces of these receptors are connected through a channel of highly conserved polar amino acids whose dynamic motions are in high correlation in the agonist- and G protein-bound active states. This and independent literature data led us to hypothesize that, in addition to consecutive conformational transitions, a shift of macroscopic polarization takes place in the transmembrane domain, which is furnished by the rearrangement of polar species through their concerted movements. Here, we examined the CB1 receptor signaling complexes utilizing microsecond scale, all-atom molecular dynamics (MD) simulations in order to see if our previous assumptions could be applied to the CB1 receptor too. Apart from the identification of the previously proposed general features of the activation mechanism, several specific properties of the CB1 have been indicated that could possibly be associated with the signaling profile of this receptor.

Isolation, characterization and in silico study of propenamide alkaloids from Hymenoepmecis bicolor poison against active μ-opioid receptor

Some insects produce venoms to defend against predators and directly interact with opioid receptors. In the present study, it was identified two alkaloids in the wasp venom species Hymenoepimecis bicolor. It was demonstrated that these could act as potential inhibitors of opioid receptors through their robust affinity to the receptors. The interaction profile was given to opioid receptors (μOR), with 60% of targets similar to alkaloid 1, with 0.25 probability, and 46.7% of targets similar to alkaloid 2, with a probability 0.17 of affinity as a target, which is considered signaling macromolecules and can mediate the most potent analgesic and addictive properties of opiate alkaloids. Notably, both alkaloids showed −7.6 kcal/mol affinity to the morphine agonies through six residues, Gly124, Asp147, Trp293, Ile296, Ile322, and Tyr326. These observations suggest further research on opioid receptors using in vitro studies of possible therapeutic applications. Communicated by Ramaswamy H. Sarma

Solid-Phase Synthesis of the Bicyclic Peptide OL-CTOP Containing Two Disulfide Bridges, and an Assessment of Its In Vivo μ-Opioid Receptor Antagonism after Nasal Administration.

New strategies facilitate the design of cyclic peptides which can penetrate the brain. We have designed a bicyclic peptide, OL-CTOP, composed of the sequences of a selective μ-opioid receptor antagonist, CTOP (f-cyclo(CYwOTX)T) (X = penicillamine, Pen; O = ornithine) and odorranalectin, OL (YASPK-cyclo(CFRYPNGVLAC)T), optimized its solid-phase synthesis and demonstrated its ability for nose-to-brain delivery and in vivo activity. The differences in reactivity of Cys and Pen thiol groups protected with trityl and/or acetamidomethyl protecting groups toward I2 in different solvents were exploited for selective disulfide bond formation on the solid phase. Both the single step and the sequential strategy applied to macrocyclization reactions generated the desired OL-CTOP, with the sequential strategy yielding a large quantity and better purity of crude OL-CTOP. Importantly, intranasally (i.n.s.) administered OL-CTOP dose-dependently antagonized the analgesic effect of morphine administered to mice through the intracerebroventricular route and prevented morphine-induced respiratory depression. In summary, the results demonstrate the feasibility of our solid-phase synthetic strategy for the preparation of the OL-CTOP bicyclic peptide containing two disulfide bonds and reveal the potential of odorranalectin for further modifications and the targeted delivery to the brain.

Altered Membrane Expression and Function of CD11b Play a Role in the Immunosuppressive Effects of Morphine on Macrophages at the Nanomolar Level.

Morphine, one of the most efficacious analgesics, is effective in severe pain, especially in patients with concomitant painful cancers. The clinical use of morphine may be accompanied by increased immunosuppression, susceptibility to infection and postoperative tumor metastatic recurrence, and the specific mechanisms and clinical strategies to alleviate this suppression remain to be investigated. Expression of CD11b is closely associated with the macrophage phagocytosis of xenobiotic particles, bacteria or tumor cells. Here, we find that morphine at 0.1-10 nM levels inhibited CD11b expression and function on macrophages via a μ-opioid receptor (MOR)-dependent mechanism, thereby reducing macrophage phagocytosis of tumor cells, a process that can be reversed by thymopentin (TP5), a commonly used immune-enhancing adjuvant in clinical practice. By knocking down or overexpressing MOR on macrophages and using naloxone, an antagonist of the MOR receptor, and LA1, a molecule that promotes macrophage CD11b activation, we suggest that morphine may regulate macrophage phagocytosis by inhibiting the surface expression and function of macrophage CD11b through the membrane expression and activation of MOR. The CD47/SIRPα axis, which is engaged in macrophage-tumor immune escape, was not significantly affected by morphine. Notably, TP5, when combined with morphine, reversed the inhibition of macrophage phagocytosis by morphine through mechanisms that promote membrane expression of CD11b and modulate its downstream signaling (e.g., NOS2, IFNG, IL1B and TNFA, as well as AGR1, PDGFB, IL6, STAT3, and MYC). Thus, altered membrane expression and function of CD11b may mediate the inhibition of macrophage phagocytosis by therapeutic doses of morphine, and the reversal of this process by TP5 may provide an effective palliative option for clinical immunosuppression by morphine.

Managing the adverse effects of cytotoxic chemotherapy at the level of primary healthcare

Background: Despite the increased incidence of cancer, more effective cancer drugs are being developed and cancer patient survival expectancy has been prolonged. Here, we discuss the contribution of primary healthcare clinics to managing the side effects of cytotoxic chemotherapy and provide a compilation of what is currently being used in this field.Current Concepts: Loperamide is recommended for active use when diarrhea occurs. In severe cases, octreotide can help control diarrhea by reducing small bowel fluid secretion. Mouthwash mixed with salt and sodium bicarbonate is helpful for controlling mucositis, and transdermal fentanyl patches can be used for severe pain. Limited evidence suggests that pilocarpine may be helpful for xerostomia. Neurokinin-1 receptor antagonists, serotonin receptor antagonists, glucocorticoids, metoclopramide, and olanzapine are used for nausea and vomiting. Opioid-related constipation can be relieved without counteracting the analgesic effect of opioids via peripherally acting μ-opioid receptor antagonists. Granulocyte colony-stimulating factor is recommended to treat neutropenic fever. In cases with neutropenia but no fever, the decision to use it should be based on the patient’s condition. Minoxidil can be used for hair loss, and scalp cooling can also be considered, but its practical applications are limited. Topical steroid ointments are used to treat hand-foot syndrome. Selective serotonin re uptake inhibitors are recommended for depression in cancer patients because they have little interaction with other drugs and are well tolerated.Discussion and Conclusion: Due to an increased cancer patient management burden and limited medical resources, primary healthcare practitioners should know these management strategies for cancer patients receiving cytotoxic chemotherapy.

Coordinated activity of a central pathway drives associative opioid analgesic tolerance.

Opioid analgesic tolerance, a root cause of opioid overdose and misuse, can develop through an associative learning. Despite intensive research, the locus and central pathway subserving the associative opioid analgesic tolerance (AOAT) remains unclear. Using a combination of chemo/optogenetic manipulation with calcium imaging and slice physiology, here we identify neuronal ensembles in a hierarchically organized pathway essential for AOAT. The association of morphine-induced analgesia with an environmental condition drives glutamatergic signaling from ventral hippocampus (vHPC) to dorsomedial prefrontal cortex (dmPFC) cholecystokininergic (CCKergic) neurons. Excitation of CCKergic neurons, which project and release CCK to basolateral amygdala (BLA) glutamatergic neurons, relays AOAT signal through inhibition of BLA μ-opioid receptor function, thereby leading to further loss of morphine analgesic efficacy. This work provides evidence for a circuit across different brain regions distinct for opioid analgesic tolerance. The components of this pathway are potential targets to treat opioid overdose and abuse.

The effect of intraperitoneal and intra-RMTg infusions of CTAP on rats’ social interaction

Social interaction is necessary for the development of individuals and society. Social interaction behaviors are rewarding. Similar to exogenous opioids, social interaction behaviors are able to induce rewarding effects that are regulated by the endogenous opioid system as well. As one type of opioid receptor, μ-opioid receptors (MORs), are densely expressed in the rostromedial tegmental nucleus (RMTg), which results in the RMTg being extremely sensitive to rewarding effects induced by exogenous and endogenous opioids. Here, we investigated how RMTg MORs played a role in rewarding effects induced by social interaction behaviors of male Wistar rats, using a conditioned place preference (CPP) model. Results showed that the CPP induced by social interaction behaviors was inhibited when the function of MORs was blocked via injecting CTAP (a selective MOR antagonist) intraperitoneally, and intra-RMTg injections of lower doses of CTAP affected the CPP in the same way. In addition, injecting CTAP intraperitoneally significantly inhibited the expression of pouncing behavior, while intra-RMTg injections of CTAP significantly inhibited the expression of all three types of social behaviors. These results suggest that RMTg MORs may be a crucial target and remain to be further explored in order to better understand the mechanism of the rewarding effects of social interaction behaviors.

Inhibitory Effects of a Novel μ-Opioid Receptor Nonpeptide Antagonist, UD-030, on Morphine-Induced Conditioned Place Preference

Although opioids are widely used to treat moderate to severe pain, opioid addiction and the opioid overdose epidemic are becoming more serious. Although opioid receptor antagonists/partial agonists, such as naltrexone and buprenorphine, have relatively low selectivity for the μ-opioid receptor (MOP), they have been used for the management of opioid use disorder. The utility of highly selective MOP antagonists remains to be evaluated. Here, we biologically and pharmacologically evaluated a novel nonpeptide ligand, UD-030, as a selective MOP antagonist. UD-030 had more than 100-fold higher binding affinity for the human MOP (Ki = 3.1 nM) than for δ-opioid, κ-opioid, and nociceptin receptors (Ki = 1800, 460, and 1800 nM, respectively) in competitive binding assays. The [35S]-GTPγS binding assay showed that UD-030 acts as a selective MOP full antagonist. The oral administration of UD-030 dose-dependently suppressed the acquisition and expression of morphine-induced conditioned place preference in C57BL/6J mice, and its effects were comparable to naltrexone. These results indicate the UD-030 may be a new candidate for the treatment of opioid use disorder, with characteristics that differ from traditional medications that are in clinical use.