Huiping Ding

A novel orvinol analog, BU08028, as a safe opioid analgesic without abuse liability in primates

Significance A potent opioid analgesic without addictive and respiratory adverse effects has been a predominant goal for opioid medicinal chemistry since the isolation of morphine from opium in the 19th century. Here we report a functional profile of a unique analog, BU08028, targeting a combination of a classical and nonclassical opioid receptors in monkeys. By examining behavioral, physiological, and pharmacologic factors, the present study demonstrates that BU08028 exhibits full antinociception and antihypersensitivity without reinforcing effects (i.e., abuse liability), respiratory depression, pruritus, adverse cardiovascular events, or acute physical dependence. Because monkey models provide the most phylogenetically appropriate evaluation of opioid receptor functions and drug effects, these findings provide a translational bridge for such ligands as effective analgesics without safety and abuse liability concerns. Despite the critical need, no previous research has substantiated safe opioid analgesics without abuse liability in primates. Recent advances in medicinal chemistry have led to the development of ligands with mixed mu opioid peptide (MOP)/nociceptin-orphanin FQ peptide (NOP) receptor agonist activity to achieve this objective. BU08028 is a novel orvinol analog that displays a similar binding profile to buprenorphine with improved affinity and efficacy at NOP receptors. The aim of this preclinical study was to establish the functional profile of BU08028 in monkeys using clinically used MOP receptor agonists for side-by-side comparisons in various well-honed behavioral and physiological assays. Systemic BU08028 (0.001–0.01 mg/kg) produced potent long-lasting (i.e., >24 h) antinociceptive and antiallodynic effects, which were blocked by MOP or NOP receptor antagonists. More importantly, the reinforcing strength of BU08028 was significantly lower than that of cocaine, remifentanil, or buprenorphine in monkeys responding under a progressive-ratio schedule of drug self-administration. Unlike MOP receptor agonists, BU08028 at antinociceptive doses and ∼10- to 30-fold higher doses did not cause respiratory depression or cardiovascular adverse events as measured by telemetry devices. After repeated administration, the monkeys developed acute physical dependence on morphine, as manifested by precipitated withdrawal signs, such as increased respiratory rate, heart rate, and blood pressure. In contrast, monkeys did not show physical dependence on BU08028. These in vivo findings in primates not only document the efficacy and tolerability profile of bifunctional MOP/NOP receptor agonists, but also provide a means of translating such ligands into therapies as safe and potentially abuse-free opioid analgesics.

Supraspinal actions of nociceptin/orphanin FQ, morphine and substance P in regulating pain and itch in non‐human primates

Nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor agonists display a promising analgesic profile in preclinical studies. However, supraspinal N/OFQ produced hyperalgesia in rodents and such effects have not been addressed in primates. Thus, the aim of this study was to investigate the effects of centrally administered ligands on regulating pain and itch in non‐human primates. In particular, nociceptive thresholds affected by intracisternal N/OFQ were compared with those of morphine and substance P, known to provide analgesia and mediate hyperalgesia, respectively, in humans.

Central N/OFQ-NOP Receptor System in Pain Modulation.

Two decades have passed since the peptide, nociceptin/orphanin FQ (N/OFQ), and its cognate (NOP) receptor were discovered. Although NOP receptor activation causes a similar pattern of intracellular actions as mu-opioid (MOP) receptors, NOP receptor-mediated pain modulation in rodents are more complicated than MOP receptor activation. This review highlights the functional evidence of spinal, supraspinal, and systemic actions of NOP receptor agonists for regulating pain. In rodents, effects of the N/OFQ-NOP receptor system in spinal and supraspinal sites for modulating pain are bidirectional depending on the doses, assays, and pain modalities. The net effect of systemically administered NOP receptor agonists may depend on relative contribution of spinal and supraspinal actions of the N/OFQ-NOP receptor signaling in rodents under different pain states. In stark contrast, NOP receptor agonists produce only antinociception and antihypersensitivity in spinal and supraspinal regions of nonhuman primates regardless of doses and assays. More importantly, NOP receptor agonists and a few bifunctional NOP/MOP receptor agonists do not exhibit reinforcing effects (abuse liability), respiratory depression, itch pruritus, nor do they delay the gastrointestinal transit function (constipation) in nonhuman primates. Depending upon their intrinsic efficacies for activating NOP and MOP receptors, bifunctional NOP/MOP receptor agonists warrant additional investigation in primates regarding their side effect profiles. Nevertheless, NOP receptor-related agonists display a much wider therapeutic window as compared to that of MOP receptor agonists in primates. Both selective NOP receptor agonists and bifunctional NOP/MOP receptor agonists hold great potential as effective and safe analgesics without typical opioid-associated side effects in humans.

Spinal antinociceptive effects of the novel NOP receptor agonist PWT2-nociceptin/orphanin FQ in mice and monkeys

Using an innovative chemical approach, peptide welding technology (PWT), a tetrabranched derivative of nociceptin/orphanin FQ (N/OFQ) has been generated and pharmacologically characterized. Both in vitro and in vivo PWT2‐N/OFQ displayed the same pharmacological profile to the natural ligand. It was more potent and produced longer‐lasting effects. The aim of the present study was to investigate the spinal effects of PWT2‐N/OFQ in nociceptive and neuropathic pain models in mice and non‐human primates.

Spinal Functions of B-Type Natriuretic Peptide, Gastrin-Releasing Peptide, and Their Cognate Receptors for Regulating Itch in Mice

B-type natriuretic peptide (BNP)–natriuretic peptide receptor A (NPRA) and gastrin-releasing peptide (GRP)–GRP receptor (GRPR) systems contribute to spinal processing of itch. However, pharmacological and anatomic evidence of these two spinal ligand-receptor systems are still not clear. The aim of this study was to determine the spinal functions of BNP-NPRA and GRP-GRPR systems for regulating scratching activities in mice by using pharmacological and immunohistochemical approaches. Our results showed that intrathecal administration of BNP (0.3–3 nmol) dose dependently elicited scratching responses, which could be blocked by the NPRA antagonist (Arg6,β-cyclohexyl-Ala8,D-Tic16,Arg17,Cys18)-atrial natriuretic factor(6-18) amide (A71915). However, A71915 had no effect on intrathecal GRP-induced scratching. In contrast, pretreatment with a GRPR antagonist (D-Tpi6,Leu13ψ(CH2-NH)-Leu14)bombesin(6-14) (RC-3095) inhibited BNP-induced scratching. Immunostaining revealed that NPRA proteins colocalize with GRP, but not GRPR, in the superficial area of dorsal horn, whereas BNP proteins do not colocalize with either GRP or GRPR in the dorsal horn. Intradermal administration of ligands including endothelin-1, U-46619, bovine adrenal medulla 8-22, and Ser-Leu-Ile-Gly-Arg-Leu-NH2 (SLIGRL) increased scratching bouts at different levels of magnitude. Pretreatment with intrathecal A71915 did not affect scratching responses elicited by all four pruritogens, whereas pretreatment with RC-3095 only inhibited SLIGRL-induced scratching. Interestingly, immunostaining showed that RC-3095, but not A71915, inhibited SLIGRL-elicited c-Fos activation in the spinal dorsal horn, which was in line with behavioral outcomes. These findings demonstrate that: 1) BNP-NPRA system may function upstream of the GRP-GRPR system to regulate itch in the mouse spinal cord, and 2) both NPRA and GRPR antagonists may have antipruritic efficacy against centrally, but not peripherally, elicited itch.

Pharmacological studies on the NOP and opioid receptor agonist PWT2-[Dmt1]N/OFQ(1-13)

&NA; An innovative chemical strategy named peptide welding technology (PWT) has been developed for the facile synthesis of tetrabranched peptides. [Dmt1]N/OFQ(1–13)‐NH2 acts as a universal agonist for nociceptin/orphanin FQ (N/OFQ) and classical opioid receptors. The present study investigated the pharmacological profile of the PWT derivative of [Dmt1]N/OFQ(1–13)NH2 (PWT2‐[Dmt1]) in several assays in vitro and in vivo after spinal administration in monkeys subjected to the tail withdrawal assay. PWT2‐[Dmt1] mimicked the effects of [Dmt1]N/OFQ(1–13)‐NH2 displaying full agonist activity, similar affinity/potency and selectivity at human recombinant N/OFQ (NOP) and opioid receptors in receptor binding, stimulation of [35S]GTP&ggr;S binding, calcium mobilization in cells expressing chimeric G proteins, and BRET studies for measuring receptor/G‐protein and receptor/&bgr;‐arrestin 2 interaction. In vivo in monkeys PWT2‐[Dmt1] elicited dose‐dependent and robust antinociceptive effects being more potent and longer lasting than [Dmt1]N/OFQ(1–13)‐NH2. The analgesic action of PWT2‐[Dmt1] was sensitive to the NOP receptor antagonist J‐113397, but not naltrexone. Thus, the present study demonstrated that the tetrabranched derivative of [Dmt1]N/OFQ(1–13)‐NH2 obtained with the PWT technology maintains the in vitro pharmacological profile of the parent peptide but displays higher potency and longer lasting action in vivo. Graphical abstract Figure. No caption available.

Altered expression of glial markers, chemokines, and opioid receptors in the spinal cord of type 2 diabetic monkeys

Neuroinflammation is a pathological condition that underlies diabetes and affects sensory processing. Given the high prevalence of pain in diabetic patients and crosstalk between chemokines and opioids, it is pivotal to know whether neuroinflammation-associated mediators are dysregulated in the central nervous system of diabetic primates. Therefore, the aim of this study was to investigate whether mRNA expression levels of glial markers, chemokines, and opioid receptors are altered in the spinal cord and thalamus of naturally occurring type 2 diabetic monkeys (n=7) compared with age-matched non-diabetic monkeys (n=6). By using RT-qPCR, we found that mRNA expression levels of both GFAP and IBA1 were up-regulated in the spinal dorsal horn (SDH) of diabetic monkeys compared with non-diabetic monkeys. Among all chemokines, expression levels of three chemokine ligand-receptor systems, i.e., CCL2-CCR2, CCL3-CCR1/5, and CCL4-CCR5, were up-regulated in the SDH of diabetic monkeys. Moreover, in the SDH, seven additional chemokine receptors, i.e., CCR4, CCR6, CCR8, CCR10, CXCR3, CXCR5, and CXCR6, were also up-regulated in diabetic monkeys. In contrast, expression levels of MOP, KOP, and DOP, but not NOP receptors, were down-regulated in the SDH of diabetic monkeys, and the thalamus had fewer changes in the glial markers, chemokines and opioids. These findings indicate that neuroinflammation, manifested as glial activation and simultaneous up-regulation of multiple chemokine ligands and receptors, seems to be permanent in type 2 diabetic monkeys. As chemokines and opioids are important pain modulators, this first-in-primate study provides a translational bridge for determining the functional efficacy of spinal drugs targeting their signaling cascades.

A bifunctional nociceptin and mu opioid receptor agonist is analgesic without opioid side effects in nonhuman primates

The small-molecule AT-121 is an agonist of nociceptin and mu opioid peptide receptors and mediates analgesia without opioid-associated side effects in nonhuman primates. A dual-targeting painkiller Opioids are among the most effective treatments for severe pain. Their pain-relieving effects are mediated by activation of the mu opioid peptide (MOP) receptor. Unfortunately, selective MOP agonists induce diverse side effects, including respiratory depression, tolerance, hyperalgesia, and dependence. Recently, activation of the nociceptin/orphanin FQ peptide (NOP) receptor has been reported to enhance MOP agonist–induced analgesia without producing side effects. Now, Ding et al. have developed a bifunctional MOP/NOP agonist, called AT-121, that showed potent analgesic effects in nonhuman primates without inducing hyperalgesia, respiratory depression, or dependence. The results suggest that bifunctional MOP/NOP agonists might represent a safe and effective pharmacological tool for treating severe pain. Misuse of prescription opioids, opioid addiction, and overdose underscore the urgent need for developing addiction-free effective medications for treating severe pain. Mu opioid peptide (MOP) receptor agonists provide very effective pain relief. However, severe side effects limit their use in the clinical setting. Agonists of the nociceptin/orphanin FQ peptide (NOP) receptor have been shown to modulate the antinociceptive and reinforcing effects of MOP agonists. We report the discovery and development of a bifunctional NOP/MOP receptor agonist, AT-121, which has partial agonist activity at both NOP and MOP receptors. AT-121 suppressed oxycodone’s reinforcing effects and exerted morphine-like analgesic effects in nonhuman primates. AT-121 treatment did not induce side effects commonly associated with opioids, such as respiratory depression, abuse potential, opioid-induced hyperalgesia, and physical dependence. Our results in nonhuman primates suggest that bifunctional NOP/MOP agonists with the appropriate balance of NOP and MOP agonist activity may provide a dual therapeutic action for safe and effective pain relief and treating prescription opioid abuse.

Differential mRNA expression of neuroinflammatory modulators in the spinal cord and thalamus of type 2 diabetic monkeys: Neuroinflammation in type 2 diabetic monkeys

Given that diabetes‐associated complications are closely associated with neuroinflammation, it is imperative to study potential changes in neuroinflammatory modulators in the central nervous system of diabetic primates.