Antinociceptive Efficacy of the µ-Opioid/Nociceptin Peptide-Based Hybrid KGNOP1 in Inflammatory Pain without Rewarding Effects in Mice: An Experimental Assessment and Molecular Docking

Abstract

Opioids are the most effective analgesics, with most clinically available opioids being agonists to the µ-opioid receptor (MOR). The MOR is also responsible for their unwanted effects, including reward and opioid misuse leading to the current public health crisis. The imperative need for safer, non-addictive pain therapies drives the search for novel leads and new treatment strategies. In this study, the recently discovered MOR/nociceptin (NOP) receptor peptide hybrid KGNOP1 (H-Dmt-D-Arg-Aba-β-Ala-Arg-Tyr-Tyr-Arg-Ile-Lys-NH2) was evaluated following subcutaneous administration in mouse models of acute (formalin test) and chronic inflammatory pain (Complete Freund’s adjuvant-induced paw hyperalgesia), liabilities of spontaneous locomotion, conditioned place preference, and the withdrawal syndrome. KGNOP1 demonstrated dose-dependent antinociceptive effects in the formalin test, and efficacy in attenuating thermal hyperalgesia with prolonged duration of action. Antinociceptive effects of KGNOP1 were reversed by naltrexone and SB-612111, indicating the involvement of both MOR and NOP receptor agonism. In comparison with morphine, KGNOP1 was more potent and effective in mouse models of inflammatory pain. Unlike morphine, KGNOP1 displayed reduced detrimental liabilities, as no locomotor impairment nor rewarding and withdrawal effects were observed. Docking of KGNOP1 to the MOR and NOP receptors and subsequent 3D interaction pattern analyses provided valuable insights into its binding mode. The mixed MOR/NOP receptor peptide KGNOP1 holds promise in the effort to develop new analgesics for the treatment of various pain states with fewer MOR-mediated side effects, particularly abuse and dependence liabilities.

Role of Nociceptin/Orphanin FQ-NOP Receptor System in the Regulation of Stress-Related Disorders

“…Hence, drugs that simultaneously activate NOP in addition to the canonical opioid receptors (especially MOP), have the potential to act as an analgesic with moderate abuse liability. Based on this hypothesis, research programs aimed at characterizing the pharmacological properties of mixed MOP/NOP agonists have attracted attention [ 43 , 44 ]. Two drugs with this mixed pharmacological profile are buprenorphine and cebranopadol.…”

Mechanistic Characterization of the Pharmacological Profile of HS-731, a Peripherally Acting Opioid Analgesic, at the µ-, δ-, κ-Opioid and Nociceptin Receptors

“…Cells were maintained in a humidified atmosphere of 95% air and 5% CO 2 . Membranes from CHO-hNOP cells were prepared as described previously [62]. Briefly, CHO-hNOP cells grown at confluence were removed from the culture plates by scraping, homogenized in 50 mM Tris-HCl buffer (pH 7.7) using a Polytron homogenizer, then centrifuged once and washed by an additional centrifugation at 27,000× g for 15 min at 4 • C. The final pellet was resuspended in 50 mM Tris-HCl buffer (pH 7.7) and stored at -80 • C until use.…”

“…Competitive binding assays at the human NOP receptor stably transfected into CHO cells were performed according to the published procedure [62]. Cell membranes (15 µg) were incubated in 50 mM Tris-HCl buffer (pH 7.4) with [ 3 H]nociceptin (0.1 nM) and various concentrations of test compounds in a final volume of 1 mL, for 60 min at 25 • C. Nonspecific binding was determined using 10 µM of unlabeled nociceptin.…”

In Vitro, In Vivo and In Silico Characterization of a Novel Kappa-Opioid Receptor Antagonist

“…Based on the in vitro results, the KOR antagonist activity of Compound A was evaluated in vivo in mouse models of visceral pain (acetic acid-induced writhing assay) and inflammatory pain (the formalin test), according to previously described procedures [51,52]. To this aim, Compound A, administered to mice subcutaneously (s.c.), was assessed for its capability to antagonize the antinociceptive effect produced by the typical KOR agonist U50,488 in both pain models (Figure 4).…”