Functional genetics reveals the contribution of delta opioid receptor to type 2 diabetes and beta-cell function.

Functional genetics has identified drug targets for metabolic disorders. Opioid use impacts metabolic homeostasis, although mechanisms remain elusive. Here, we explore the OPRD1 gene (encoding delta opioid receptor, DOP) to understand its impact on type 2 diabetes. Large-scale sequencing of OPRD1 and in vitro analysis reveal that loss-of-function variants are associated with higher adiposity and lower hyperglycemia risk, whereas gain-of-function variants are associated with lower adiposity and higher type 2 diabetes risk. These findings align with studies of opium addicts. OPRD1 is expressed in human islets and beta cells, with decreased expression under type 2 diabetes conditions. DOP inhibition by an antagonist enhances insulin secretion from human beta cells and islets. RNA-sequencing identifies pathways regulated by DOP antagonism, including nerve growth factor, circadian clock, and nuclear receptor pathways. Our study highlights DOP as a key player between opioids and metabolic homeostasis, suggesting its potential as a therapeutic target for type 2 diabetes.

Identification of Cytisine Derivatives as Agonists of the Human Delta Opioid Receptor by Supercomputer-Based Virtual Drug Screening and Transcriptomics.

Delta opioid receptors (DORs) are rising as therapeutic targets, not only for the treatment of pain but also other neurological disorders (e.g., Parkinson's disease). The advantage of DOR agonists compared to µ-opioid receptor agonists is that they have fewer side effects and a lower potential to induce tolerance. However, although multiple candidates have been tested in the past few decades, none have been approved for clinical use. The current study focused on searching for new DOR agonists by screening a chemical library containing 40,000 natural and natural-derived products. The functional activity of the top molecules was evaluated in vitro through the cyclic adenosine monophosphate accumulation assay. Compound 3 showed promising results, and its activity was further investigated through transcriptomic methods. Compound 3 inhibited the expression of TNF-α, prevented NF-κB translocation to the nucleus, and activated the G-protein-mediated ERK1/2 pathway. Additionally, compound 3 is structurally different from known DOR agonists, making it a valuable candidate for further investigation for its anti-inflammatory and analgesic potential.

Design, Synthesis, and Characterization of New δ Opioid Receptor-Selective Fluorescent Probes and Applications in Single-Molecule Microscopy of Wild-Type Receptors.

The delta opioid receptor (δOR or DOR) is a G protein-coupled receptor (GPCR) showing a promising profile as a drug target for nociception and analgesia. Herein, we design and synthesize new fluorescent antagonist probes with high δOR selectivity that are ideally suited for single-molecule microscopy (SMM) applications in unmodified, untagged receptors. Using our new probes, we investigated wild-type δOR localization and mobility at low physiological receptor densities for the first time. Furthermore, we investigate the potential formation of δOR homodimers, as such a receptor organization might exhibit distinct pharmacological activity, potentially paving the way for innovative pharmacological therapies. Our findings indicate that the majority of δORs labeled with these probes exist as freely diffusing monomers on the cell surface in a simple cell model. This discovery advances our understanding of OR behavior and offers potential implications for future therapeutic research.

Structure-Activity Relationship Study of CYM51010, an agonist for the µ-δ Opioid Receptor Heterodimer.

Although opioid analgesics are indispensable in treating pain, these drugs are accompanied by life-threatening side effects. While clinically relevant opioid drugs target the µ opioid receptor (MOR), a heterodimer between the MOR and the δ opioid receptor (DOR) has emerged as another target to develop safer analgesics. Although some heterodimer-preferring agonists have been reported so far, it is still difficult to activate the MOR/DOR heterodimer selectively in the presence of MOR or DOR monomers/homodimers. To gain insights to develop selective agonists for MOR/DOR, herein we prepared analogs of CYM51010, one of the reported heterodimer-preferring agonists, and collected structure-activity relationship information. We found that the ethoxycarbonyl group was needed for the activity for the heterodimer, although this group could be substituted with functional groups with similar sizes, such as an ethoxycarbonyl group. As for the acetylaminophenyl group, not a type of substituent, but rather a substituent located at a specific position (para-position) was essential for the activity. Changing the linker length between the acetylaminophenyl group and the piperidine moiety also had deleterious effects on the activity. On the other hand, the substitution of the acetylamino group with a trifluoroacetylamino group and the substitution of the phenethyl group with a benzyl group diminished the activities for the monomers/homodimers while keeping the activity for MOR/DOR, which enhanced the selectivity. Our findings herein will play an important role in developing selective agonists for MOR/DOR and for elucidating the physiological roles of this heterodimer in analgesic processes and in the establishment of side effects.

Effects of Selective and Mixed-Action Kappa and Delta Opioid Receptor Agonists on Pain-Related Behavioral Depression in Mice.

We recently developed a series of nalfurafine analogs (TK10, TK33, and TK35) that may serve as non-addictive candidate analgesics. These compounds are mixed-action agonists at the kappa and delta opioid receptors (KOR and DOR, respectively) and produce antinociception in a mouse warm-water tail-immersion test while failing to produce typical mu opioid receptor (MOR)-mediated side effects. The warm-water tail-immersion test is an assay of pain-stimulated behavior vulnerable to false-positive analgesic-like effects by drugs that produce motor impairment. Accordingly, this study evaluated TK10, TK33, and TK35 in a recently validated assay of pain-related behavioral depression in mice that are less vulnerable to false-positive effects. For comparison, we also evaluated the effects of the MOR agonist/analgesic hydrocodone (positive control), the neurokinin 1 receptor (NK1R) antagonist aprepitant (negative control), nalfurafine as a selective KOR agonist, SNC80 as a selective DOR agonist, and a nalfurafine/SNC80 mixture. Intraperitoneal injection of dilute lactic acid (IP lactic acid) served as a noxious stimulus to depress vertical and horizontal locomotor activity in male and female ICR mice. IP lactic acid-induced locomotor depression was alleviated by hydrocodone but not by aprepitant, nalfurafine, SNC80, the nalfurafine/SNC80 mixture, or the KOR/DOR agonists. These results suggest that caution is warranted in advancing mixed-action KOR/DOR agonists as candidate analgesics.

Descriptive molecular pharmacology of the δ opioid receptor (DOR): A computational study with structural approach.

This work focuses on the δ receptor (DOR), a G protein-coupled receptor (GPCR) belonging to the opioid receptor group. DOR is expressed in numerous tissues, particularly within the nervous system. Our study explores computationally the receptor's interactions with various ligands, including opiates and opioid peptides. It elucidates how these interactions influence the δ receptor response, relevant in a wide range of health and pathological processes. Thus, our investigation aims to explore the significance of DOR as an incoming drug target for pain relief and neurodegenerative diseases and as a source for novel opioid non-narcotic analgesic alternatives. We analyze the receptor's structural properties and interactions using Molecular Dynamics (MD) simulations and Gaussian-accelerated MD across different functional states. To thoroughly assess the primary differences in the structural and conformational ensembles across our different simulated systems, we initiated our study with 1 µs of conventional Molecular Dynamics. The strategy was chosen to encompass the full activation cycle of GPCRs, as activation processes typically occur within this microsecond range. Following the cMD, we extended our study with an additional 100 ns of Gaussian accelerated Molecular Dynamics (GaMD) to enhance the sampling of conformational states. This simulation approach allowed us to capture a comprehensive range of dynamic interactions and conformational changes that are crucial for GPCR activation as influenced by different ligands. Our study includes comparing agonist and antagonist complexes to uncover the collective patterns of their functional states, regarding activation, blocking, and inactivation of DOR, starting from experimental data. In addition, we also explored interactions between agonist and antagonist molecules from opiate and opioid classifications to establish robust structure-activity relationships. These interactions have been systematically quantified using a Quantitative Structure-Activity Relationships (QSAR) model. This research significantly contributes to our understanding of this significant pharmacological target, which is emerging as an attractive subject for drug development.

In-silico screening of missense nsSNPs in Delta-opioid receptor protein and their restoring tendency on MCRT interaction; focusing on dynamic nature.

Delta-opioid receptor protein (OPRD1) is one of the potential targets for treating pain. The presently available opioid agonists are known to cause unnecessary side effects. To discover a novel opioid agonist, our research group has synthesized a chimeric peptide MCRT and proved its potential activity through in vivo analysis. Non-synonymous SNPs (nsSNPs) missense mutations affect the functionality and stability of proteins leading to diseases. The current research was focused on understanding the role of MCRT in restoring the binding tendency of OPRD1 nsSNPs missense mutations on dynamic nature in comparison with Deltorphin-II and morphiceptin. The deleterious effects of nsSNPs were analyzed using various bioinformatics tools for predicting structural, functional, and oncogenic influence. The shortlisted nine nsSNPs were predicted for allergic reactions, domain changes, post-translation modification, multiple sequence alignment, secondary structure, molecular dynamic simulation (MDS), and peptide docking influence. Further, the docked complex of three shortlisted deleterious nsSNPs was analyzed using an MDS study, and the highly deleterious shortlisted nsSNP A149T was further analyzed for higher trajectory analysis. MCRT restored the binding tendency influence caused by nsSNPs on the dynamics of stability, functionality, binding affinity, secondary structure, residues connection, motion, and folding of OPRD1 protein.

Navacaprant, a novel and selective kappa opioid receptor antagonist, has no agonist properties implicated in opioid-related abuse.

Kappa opioid receptors (KORs) are implicated in the pathophysiology of various psychiatric and neurological disorders creating interest in targeting the KOR system for therapeutic purposes. Accordingly, navacaprant (NMRA-140) is a potent, selective KOR antagonist being evaluated as a treatment for major depressive disorder. In the present report, we have extended the pharmacological characterization of navacaprant by further demonstrating its selective KOR antagonist properties and confirming its lack of agonist activity at KORs and related targets involved in opioid-related abuse. Using CHO-K1 cells expressing human KOR, mu (MOR), or delta (DOR) opioid receptors, navacaprant demonstrated selective antagonist properties at KOR (IC50 = 0.029 µM) versus MOR (IC50 = 3.3 µM) and DOR (IC50 > 10 µM) in vitro. In vivo, navacaprant (10-30 mg/kg, i.p.) dose-dependently abolished KOR-agonist induced analgesia in the mouse tail-flick assay. Additionally, navacaprant (10, 30 mg/kg, p.o.) significantly reduced KOR agonist-stimulated prolactin release in mice and rats, confirming KOR antagonism in vivo. Navacaprant showed no agonist activity at any opioid receptor subtype (EC50 > 10 µM) in vitro and exhibited no analgesic effect in the tail-flick assays at doses ≤100 mg/kg, p.o. thereby confirming a lack of opioid receptor agonist activity in vivo. Importantly, navacaprant did not alter extracellular dopamine concentrations in the nucleus accumbens shell of freely-moving rats following doses ≤100 mg/kg, p.o., whereas morphine (10, 20 mg/kg, i.p.) significantly increased dopamine levels. These results demonstrate that navacaprant is a KOR-selective antagonist with no pharmacological properties implicated in opioid-related abuse.

[Underlying mechanisms for psychotropic effects of delta opioid receptor agonists].

Growing evidence has indicated that delta opioid receptor (DOP) agonists are potential psychotropic drugs such as for depression, anxiety, and PTSD. In rodent studies, we have also demonstrated that DOP agonists exhibit potent anxiolytic-like effects via the inhibition of the excitatory neuronal activity which projects to the amygdala from the prelimbic prefrontal cortex and facilitate extinction learning of contextual fear memory through PI3K-Akt signaling pathway in the infralimbic prefrontal cortex and MEK-ERK signaling pathway in the amygdala. In this article, we introduce the functional mechanisms underlying antidepressant-like effects and anti-stress effects of DOP agonists. Then, we employed a valid animal model of depression, chronic vicarious social defeat stress (cVSDS) mice, and investigated that the influence of DOP activation on pathopsychological factors in depression such as the adult hippocampal neurogenesis, hypothalamic-pituitary-adrenal (HPA) axis, and neuroinflammation. First, repeated administrations after the stress period to cVSDS mice with a selective DOP agonist, KNT-127, improved social interaction behaviors and reduced hyperactivation of the HPA axis without affecting hippocampal neurogenesis. Meanwhile, repeated KNT-127 administrations during the cVSDS period prevented the exacerbation of social interaction behaviors, dysregulation of the HPA axis, and excessive new-born neuronal cell death in the hippocampal dentate gyrus. Moreover, in both administration paradigms, KNT-127 suppressed microglial overactivation in the dentate gyrus of cVSDS mice. These results indicate that the underlying mechanism of DOP-induced antidepressant-like effects differ from those of conventional monoaminergic antidepressants. Furthermore, we propose that DOP agonists might have prophylactic effects as well as therapeutic effects on pathophysiological changes in depression.

Neuroprotective effects of miRNA-326 knockout in neonatal hypoxic-ischemic brain damage mice via the δ-opioid receptor.

Hypoxic-ischemic brain damage (HIBD) in the perinatal period is an important cause of cerebral damage and long-term neurological sequelae, and can place much pressure on families and society. Our previous study demonstrated that miRNA-326 reduces neuronal apoptosis by up-regulating the δ-opioid receptor (DOR) under oxygen-glucose deprivation in vitro. In the present study, we aimed to explore the neuroprotective effects of the miRNA-326/DOR axis by inhibiting apoptosis in HIBD using neonatal miRNA-326 knockout mice. Neonatal C57BL/6 mice, neonatal miRNA-326 knockout mice, and neonatal miRNA-326 knockout mice intraperitoneally injected with the DOR inhibitor naltrindole were treated with hypoxic-ischemia (HI). Neurological deficit scores, magnetic resonance imaging, terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end labeling, and Caspase-3, Bax, and B-cell lymphoma 2 (Bcl-2) expression were evaluated on day 2 after HI. Neurobehavioral analyses were performed on days 2 and 28 after HI. Additionally, the Morris water maze test was conducted on days 28. Compared with HI-treated neonatal C57BL/6 mice, HI-treated neonatal miRNA-326 knockout mice had higher neurological deficit scores, smaller cerebral infarction areas, and improved motor function, reaction ability, and long-term spatial learning and memory. These effects were likely the result of inhibiting apoptosis; the DOR inhibitor reversed these neuroprotective effects. Our findings indicate that miRNA-326 knockout plays a neuroprotective effect in neonatal HIBD by inhibiting apoptosis via the target gene DOR.

Alcohol consumption does not impact delta and kappa opioid receptor-mediated synaptic depression in dorsolateral striatum of adult male mice.

Many drugs of abuse, including alcohol, disrupt long-term synaptic depression (LTD) at dorsal striatal glutamate synapses. This disruption is common to many forms of LTD that are mediated by G protein coupled receptors (GPCRs) that signal through the inhibitory Gi/o class of G proteins. A loss of LTD is thought to mediate behavioral changes associated with the development of substance use disorders. We have previously shown in multiple studies that LTD mediated by the Gi/o-coupled mu opioid receptor is disrupted by in vivo opioid and alcohol exposure in adolescent and adult mice. One of our previous studies suggested that LTD mediated by delta and kappa opioid receptors was resistant to the LTD-disrupting properties of in vivo opioid exposure. We hypothesized that delta and kappa opioid receptor-mediated LTD would be exceptions to the generalizable observation that forms of dorsal striatal Gi/o-coupled receptor LTD are disrupted by drugs of abuse. Specifically, we predicted that these forms of LTD would be resistant to the deleterious effects of alcohol consumption, just as they were resistant to opioid exposure. Indeed, in adult male mice that drank alcohol for 3 weeks, delta and kappa opioid receptor-mediated LTD at glutamatergic inputs to direct pathway and indirect pathway medium spiny neurons in the dorsolateral striatum was unaffected by alcohol. These data demonstrate that alcohol effects on GPCR-mediated LTD are not generalizable across all types of Gi/o-coupled GPCRs.

Delta opioid receptor expression correlates to skin ageing and melanin expression in Asian women.

While the evidence for the implication of opioid receptors (OPr) in ageing is growing, there is, to our knowledge, no study focusing directly on changes in vivo cutaneous OPr expression with increasing age. We thus investigated OPr expression in 30 healthy female Asian volunteers in Southern China whose ages range from the early 20s to the early 60s. Excisional biopsies were taken from the sun-exposed extensor area of the lower arm and the photo-protected area of the upper inner arm. The thickness of the epidermal layers, melanin content, as well as expression of mu-opioid receptors (MOPr) and delta-opioid receptors (DOPr) were compared between different age ranges and photo-exposure status. Significant increased epidermal hypertrophy on the extensor surface was observed. There was significant reduction of DOPr in the epidermis with increasing age, independent of photo-ageing. The increase of melanin was significantly correlated with epidermal DOPr expression, not with MOPr expression. DOPr expression could thus serve as a marker for real biological ageing unaffected by chronic photo-exposure. Additionally, DOPr expression was inversely correlated with the deposition of melanin. Based on these results, we hypothesise that regulation of DOPr expression could be used to improve aged skin, including hyperpigmentation.

Activation of Cardiac δ2-Opioid Receptors Increases Heart Tolerance to Reperfusion.

Coronary occlusion (45 min) and reperfusion (120 min) in male Wistar rats in vivo, as well as total ischemia (45 min) of an isolated rat heart followed by reperfusion (30 min) were reproduced. The selective δ2-opioid receptor agonist deltorphin II (0.12 mg/kg and 152 nmol/liter) was administered intravenously 5 min before reperfusion in vivo or added to the perfusion solution at the beginning of reperfusion of the isolated heart. The peripheral opioid receptor antagonist naloxone methiodide and δ2-opioid receptor antagonist naltriben were used in doses of 5 and 0.3 mg/kg, respectively. It was found that the infarct-limiting effect of deltorphin II is associated with the activation of δ2-opioid receptors. We have demonstrated that deltorphin II can improve the recovery of the contractility of the isolated heart after total ischemia.

Expression profile and immunomodulatory roles of methionine-enkephalin and delta opioid receptor in Octopus ocellatus.

In this study, the expressions and distributions of methionine-enkephalin (Met-enk) and δ opioid receptor in the nervous system of Octopus ocellatus, and the immune regulatory mechanisms of Met-enk on O. ocellatus were explored. The distributions and expressions of Met-enk and δ opioid receptor were assessed by immunohistochemistry and enzyme-linked immunosorbent assay. UV-spectrophotometer, microplate reader, and flow cytometer were used to examine the effects of different concentrations of Met-enk on phagocytosis, antioxidant effects, and body surface mucus immunity of O. ocellatus hemocytes. The data were used to study the mechanisms of Met-enk immunity regulation in O. ocellatus. According to the results, the expression levels of Met-enk and δ opioid receptor in O. ocellatus lymphocytes were higher than those in hemocytes. The expression levels of Met-enk in the ganglia of O. ocellatus decreased in the following order: pedal ganglia > cerebral ganglia > visceral ganglia > optic ganglia > stellate ganglia. Moreover, the phagocytic activity of O. ocellatus hemocytes was enhanced with increasing Met-enk concentration. With increasing Met-enk concentration, the expressions of nitric oxide, total nitric oxide synthase, inducible nitric oxide synthase, catalase, hydrogen peroxide, myeloperoxidase, reduced glutathione, α-naphthy acetate esterase, and methionine aminopeptidases decreased in serums of O. ocellatus in the experimental group compared to the blank group. Similarly, the content of reduced glutathione in the hemocytes of O. ocellatus was also lower in the experimental group than in the blank group; however, the expressions of other substances were higher compared to the blank group. Furthermore, α-naphthy acetate esterase, myeloperoxidase, and hydrogen peroxide expressions in mucus immunity trials of the body surface were lower in the experimental group compared to the blank group. These results indicate that the distributions and expressions of Met-enk and δ opioid receptor in the nervous system of O. ocellatus were related to axoplasmic transport and immune regulation mechanisms. Met-enk participates in cellular immunity, humoral immunity, and mucus immunity in the form of neurotransmitters, thereby regulating the immune response of O. ocellatus.

Identification of 1,3,8-Triazaspiro[4.5]Decane-2,4-Dione Derivatives as a Novel δ Opioid Receptor-Selective Agonist Chemotype.

δ opioid receptors (DORs) hold potential as a target for neurologic and psychiatric disorders, yet no DOR agonist has proven efficacious in critical phase II clinical trials. The exact reasons for the failure to produce quality drug candidates for the DOR are unclear. However, it is known that certain DOR agonists can induce seizures and exhibit tachyphylaxis. Several studies have suggested that those adverse effects are more prevalent in delta agonists that share the (+)-4-[(αR)-α-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80)/4-[(αR*)-α-((2S*,5R*)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-hydroxybenzyl]-N,N-diethylbenzamide chemotype. There is a need to find novel lead candidates for drug development that have improved pharmacological properties to differentiate them from the current failed delta agonists. Our objective in this study was to identify novel DOR agonists. We used a ß-arrestin assay to screen a small G-protein coupled receptors (GPCR)-focused chemical library. We identified a novel chemotype of DOR agonists that appears to bind to the orthosteric site based of docking and molecular dynamic simulation. The most potent agonist hit compound is selective for the DOR over a panel of 167 other GPCRs, is slightly biased toward G-protein signaling and has anti-allodynic efficacy in a complete Freund's adjuvant model of inflammatory pain in C57BL/6 male and female mice. The newly discovered chemotype contrasts with molecules like SNC80 that are highly efficacious ß-arrestin recruiters and may suggest this novel class of DOR agonists could be expanded on to develop a clinical candidate drug. SIGNIFICANCE STATEMENT: δ opioid receptors are a clinical target for various neurological disorders, including migraine and chronic pain. Many of the clinically tested delta opioid agonists share a single chemotype, which carries risks during drug development. Through a small-scale high-throughput screening assay, this study identified a novel δ opioid receptor agonist chemotype, which may serve as alternative for the current analgesic clinical candidates.

The Role of Opioid Receptor Antagonists in Regulation of Blood Pressure and T-Cell Activation in Mice Selected for High Analgesia Induced by Swim Stress.

Opioid peptides and their G protein-coupled receptors are important regulators within the cardiovascular system, implicated in the modulation of both heart and vascular functions. It is known that naloxone-an opioid antagonist-may exert a hypertensive effect. Recent experimental and clinical evidence supports the important role of inflammatory mechanisms in hypertension. Since opioids may play a role in the regulation of both blood pressure and immune response, we studied these two processes in our model. We aimed to evaluate the effect of selective and non-selective opioid receptor antagonists on blood pressure and T-cell activation in a mouse model of high swim stress-induced analgesia. Blood pressure was measured before and during the infusion of opioid receptor antagonists using a non-invasive tail-cuff measurement system. To assess the activation of T-cells, flow cytometry was used. We discovered that the non-selective antagonism of the opioid system by naloxone caused a significant elevation of blood pressure. The selective antagonism of µ and κ but not δ opioid receptors significantly increased systolic blood pressure. Subsequently, a brief characterization of T-cell subsets was performed. We found that the blockade of µ and δ receptors is associated with the increased expression of CD69 on CD4 T-cells. Moreover, we observed an increase in the central memory CD4 and central memory CD8 T-cell populations after the δ opioid receptor blockade. The antagonism of the µ opioid receptor increased the CD8 effector and central memory T-cell populations.

Pharmacologic Characterization of Substituted Nitazenes at µ, κ, and Δ Opioid Receptors Suggests High Potential for Toxicity.

The benzimidazole opioids (substituted nitazenes) are highly potent µ opiod receptor (MOR) agonists with heroin- or fentanyl-like effects. These compounds have caused hospitalizations and fatal overdoses. We characterized the in vitro pharmacology and structure-activity relationships of 19 nitazenes with substitutions at three positions of the benzimidazole core. Affinities were assessed using agonist radioligand binding assays at human µ, κ, and Δ opioid receptors (MOR, KOR, and DOR, respectively) heterologously expressed in CHO cells. Notably, for MOR binding, nine substituted nitazenes had significantly higher affinities than fentanyl including N-pyrrolidino etonitazene, N-pyrrilidino isonitazene, and N-desethyl isotonitazene; 13 had subnanomolar affinities. Only metodesnitazene and flunitazene had significantly lower affinities than fentanyl. Affinities for the substituted nitazenes at KOR and DOR relative to MOR were 46- to 2580-fold and 180- to 1280-fold lower, respectively. Functional activities were assessed using [35S]GTPγS binding assays. Four nitazenes had subnanomolar potencies at MOR: N-pyrrolidino etonitazene, N-pyrrilidino isonitazene, N-pyrrilidino protonitazene and N-desethyl isotonitazene. Ten substituted nitazenes had significantly higher potencies than fentanyl. All tested nitazenes were full MOR agonists. Potencies at KOR and DOR relative to MOR were 7.3- to 7920-fold and 24- to 9400-fold lower, respectively. Thus, many of these compounds are high affinity/high potency MOR agonists with elevated potential to elicit toxicity and overdose at low doses. SIGNIFICANCE STATEMENT: Substituted nitazenes are a growing public health threat. Although the 19 nitazenes tested vary in their opioid receptor pharmacology, a number are very high affinity, high potency, and high efficacy compounds- higher than fentanyl. Their pharmacology suggests high potential for harm.

The Role of δ2-Opioid Receptors in the Regulation of Tolerance of Isolated Cardiomyocytes to Hypoxia and Reoxygenation.

Hypoxia (20 min) and reoxygenation (30 min) were simulated on isolated rat cardiomyocytes to evaluate the cytoprotective effect of selective δ2-opioid receptor agonist deltorphin II, opioid receptor antagonist naloxone methiodide, µ-opioid receptor antagonist CTAP, κ-opioid receptor antagonist nor-binaltorphimine, ε1-opioid receptor antagonist BNTX, and δ2-opioid receptors naltriben. Deltorphin II was administered 5 min before reoxygenation, antagonists were administered 10 min before reoxygenation. The cytoprotective effect of deltorphin II was assessed by the number of cardiomyocytes survived after hypoxia/reoxygenation, as well as by the lactate dehydrogenase content in the incubation medium. It has been established that the cytoprotective effect of deltorphin II occurs at a concentration of 64 nmol/liter and is associated with activation of δ2-opioid receptors.

Comparative Analysis of Infarct-Limiting Activity of Peptide and Non-Peptide δ- and κ-Opioid Receptor Agonists during Heart Reperfusion In Vivo.

A comparative analysis of the infarct-limiting activity of δ- and κ-opioid receptors (OR) agonists was carried out on a model of coronary occlusion (45 min) and reperfusion (120 min) in male Wistar rats. We used selective δ2-OR agonist deltorphin II (0.12 mg/kg), δ-OR agonists BW373U86 and p-Cl-Phe DPDPE (0.1 and 1 mg/kg), selective agonists of δ1-OR DPDPE (0.1 and 0.969 mg/kg), κ1-OR U-50,488 (0.1 and 1 mg/kg), κ2-OR GR-89696 (0.1 mg/kg), and κ-OR ICI-199,441 (0.1 mg/kg). All drugs were administered intravenously 5 min before reperfusion. Deltorphin II, BW373U86 (1 mg/kg), p-Cl-Phe DPDPE (1 mg/kg), U-50,488 (1 mg/kg), and ICI-199,441 had a cardioprotective effect. The most promising compounds for drug development are ICI-199,441 and deltorphin II.

A Novel Leu-Enkephalin Prodrug Produces Pain-Relieving and Antidepressant Effects.

Persistent pain is a significant healthcare problem with limited treatment options. The high incidence of comorbid chronic pain and depression significantly reduces life quality and complicates the treatment of both conditions. Antidepressants are less effective for pain and depression than for depression alone and they induce severe side effects. Opioids are highly efficacious analgesics, but rapid development of tolerance, dependence, and debilitating side effects limit their efficacy and safe use. Leucine-enkephalin (Leu-ENK), the endogenous delta opioid receptor agonist, controls pain and mood and produces potent analgesia with reduced adverse effects compared to conventional opioids. High proteolytic instability, however, makes Leu-ENK ineffective after systemic administration and limits its clinical usefulness. KK-103, a Leu-ENK prodrug, was developed to overcome these limitations of Leu-ENK via markedly increased plasma stability in mice. We showed rapid and substantially increased systemic adsorption and blood plasma exposure of KK-103 compared to Leu-ENK. We also observed brain uptake of radiolabeled KK-103 after systemic administration, indicating a central effect of KK-103. We then established KK-103's prolonged antinociceptive efficacy in the ramped hot plate and formalin test. In both models, KK-103 produced a comparable dose to the maximum antinociceptive-effect relationship. The pain-alleviating effect of KK-103 primarily resulted from activating the delta opioid receptor after the likely conversion of KK-103 to Leu-ENK in vivo. Finally, KK-103 produced an antidepressant-like activity comparable to the antidepressant desipramine, but with minimal gastrointestinal inhibition and no incidence of sedation.