Elsevier

Cellular Signalling

Volume 32, April 2017, Pages 59-65
Cellular Signalling

Nalfurafine is a G-protein biased agonist having significantly greater bias at the human than rodent form of the kappa opioid receptor

https://doi.org/10.1016/j.cellsig.2017.01.016Get rights and content

Highlights

  • Nalfurafine is a centrally-active G protein (ERK1/2) biased KOR agonist.

  • Nalfurafine is a potent KOR-mediated analgesic and anti-puritic.

  • Transient tolerance can be induced by high nalfurafine concentrations.

  • Nalfurafine may be a valuable structure for KOR agonist development.

Abstract

Nalfurafine is a moderately selective kappa opioid receptor (KOR) analgesic with low incidence of dysphoric side effects in clinical development for the treatment of uremic pruritis. The basis for its reduced dysphoric effect compared to other KOR agonists is not clear, but prior studies suggest that the aversive properties of KOR agonists require p38α MAPK activation through an arrestin-dependent mechanism. To determine whether nalfurafine is a functionally selective KOR agonist, we measured its potency to activate the G protein-dependent early phase of Extracellular Signal-Regulated Kinase (ERK1/2) phosphorylation and the arrestin-dependent late phase of p38 MAPK signaling. Nalfurafine was approximately 250 fold more potent for ERK1/2 activation as compared to p38 MAPK activation in human KOR (hKOR) expressing HEK293 cells, and approximately 20 fold more potent for ERK1/2 activation than p38 activation in rodent KOR (rKOR) expressing HEK293 cells. The 10-fold greater G-bias at the hKOR than rKOR was unexpected, however the G protein biased effect of nalfurafine is consistent with its reduced dysphoric effects in human and rodent models. Although nalfurafine is reported to have low receptor selectivity in radioligand binding assays, its antinociceptive effect was blocked by the selective KOR antagonist norbinaltorphimine. Nalfurafine pretreatment also resulted in a KOR-dependent and mu opioid receptor-independent reduction in scratching induced by 5′-GNTI. These findings suggest that nalfurafine is a functionally selective KOR agonist and that KOR agonists able to selectively activate G protein signaling without activating p38α MAPK may have therapeutic potential as non-dysphoric antipruritic analgesics.

Introduction

Mu opioid agonists including morphine-like opiates remain the principal analgesics for the treatment severe pain, however the respiratory depression, constipating, and addiction risks of these drugs are well known liabilities and diversion of prescription opioids is currently a major public health crisis [1]. Kappa opioid analgesics have been developed as safer alternatives to mu opioids for control of pain, however the selective kappa opioid receptor (KOR) analgesics that were initially developed produce profound dysphoric and psychotomimetic actions [2]. Attempts to reduce the dysphoric properties of KOR agonists by restricting their brain penetrance have been made; however these peripherally restricted compounds have demonstrated only modest analgesic activity [3], [4]. Recent studies have suggested that the analgesic effects of KOR are mediated by G protein regulation of ion channels, including increased G protein gated potassium channel and decreased calcium channel conductance resulting in reduced neuronal excitability; whereas the aversive effects of KOR agonists require G protein-coupled receptor kinase 3 (GRK3)-dependent arrestin activation [5], [6]. The concept that functionally selective agonists that preferentially activate one signaling cascade can be developed has re-energized opioid drug development [7], [8]. Furthermore the hypothesis that a centrally active G protein biased KOR agonist that does not induce arrestin-dependent signaling might be a safer analgesic has stimulated the field [8], [9].

Functionally selective KOR agonists have been recently identified [10], [11], [12], however these compounds lack drug-like qualities. In contrast, the moderately receptor selective KOR agonist nalfurafine (TRK820) is currently in clinical trials for the treatment of uremic pruritis [13], and has pharmacological properties suggestive of functional selectivity (e.g. analgesic, low incidence of dysphoria) [14], [15], [16]. In the present study, we assessed the functional selectivity of nalfurafine. Identification of a drug-like lead compound in the development of functionally selective KOR agonists would be a significant advance in the generation of novel analgesics potentially lacking the addictive effects of mu opioids and the dysphoric effects of conventional KOR agonists.

Section snippets

Chemicals/reagents

For cell culture experiments KOR agonists (−)U50,488 (Tocris) and nalfurafine hydrochloride ((2E)-N-[(5α,6β)-17-(cyclopropylmethyl)-3,14-dihydroxy-4,5-epoxymorphinan-6-yl]-3-(3-furyl)-N-methylacrylamide) (NIDA Drug Supply) and the MOR agonist DAMGO ([D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin) (Sigma-Aldrich) were dissolved in water. For in vivo studies, the KOR agonists nalfurafine, (+/−)U50,488, and the KOR antagonists 5′-GNTI, and norBNI were obtained from the NIDA Drug Supply and were freshly

Does nalfurafine differentially activate p38 and ERK1/2 MAPK pathways by rKOR?

KOR agonists have previously been shown to promote ERK1/2 phosphorylation at 5 min and p38 phosphorylation at 30 min via different signaling cascades [21], [26]. We first sought to establish whether nalfurafine activates these cascades with similar efficacy and potency. HEK293 cells stably expressing rKOR were treated for 5 or 30 min with nalfurafine (10 pM-100 nM) or U50,488 (1 μM) prior to lysis and phospho-ERK1/2 (5 min) or phospho-p38 (30 min) was quantified by western blot (Fig. 1A–B). Nalfurafine

Discussion

The major finding of this study is that nalfurafine is a G protein biased ligand with greater bias at the human than rodent KOR. Nalfurafine is biased towards ERK1/2 activation (G protein-dependent), with 20 × and 250 × lower potency for p38 activation by rKOR and hKOR, respectively. These findings have implications for the development of KOR agonists with therapeutic use, as p38 contributes the aversive properties of KOR agonists [5], [6], [33]. We also found that nalfurafine-induced thermal

Conclusions

Despite lacking optimal receptor selectivity, nalfurafine is a centrally-active prototype G protein biased agonist and therefore provides a base structure for future analgesic drug development. An ideal KOR agonist would be highly efficacious for G protein-mediated pathways with minimal efficacy or greatly reduced potency for GRK3 mediated-p38 activation. Additional qualities would be low affinity and minimal efficacy for other opioid receptors, notably MOR, and being centrally active. This

Funding

This work was supported by the National Institute on Drug Abuse [Grants: PO1-DA035764 and T32-DA07278].

Authorship contributions

Participated in research design: Schattauer, Kuhar, Song, Chavkin.

Conducted experiments: Schattauer, Kuhar, Song.

Performed data analysis: Schattauer, Kuhar, Song, Chavkin.

Wrote or contributed to the writing of the manuscript: Schattauer, Chavkin.

Acknowledgements

We thank Dr. John Pintar for providing the original MOR−/− and KOR−/− mice. We thank Dan Messinger for managing the mouse breeding and genotyping. We thank Dr. Peter Groblewski for technical assistance.

References (36)

  • T. Suzuki et al.

    Effect of repeated administration of TRK-820, a kappa-opioid receptor agonist, on tolerance to its antinociceptive and sedative actions

    Brain Res.

    (2004)
  • S. Inan et al.

    Investigation of gastrin-releasing peptide as a mediator for 5′-guanidinonaltrindole-induced compulsive scratching in mice

    Peptides

    (2011)
  • K.M. Dimattio et al.

    Zyklophin, a short-acting kappa opioid antagonist, induces scratching in mice

    Neurosci. Lett.

    (2014)
  • H. Nagase et al.

    Drug design and synthesis of a novel kappa opioid receptor agonist with an oxabicyclo[2.2.2] octane skeleton and its pharmacology

    Bioorg. Med. Chem. Lett.

    (2010)
  • L. Manchikanti et al.

    Opioid epidemic in the United States

    Pain Physician

    (2012)
  • L. Arendt-Nielsen et al.

    Analgesic efficacy of peripheral kappa-opioid receptor agonist CR665 compared to oxycodone in a multi-modal, multi-tissue experimental human pain model: selective effect on visceral pain

    Anesthesiology

    (2009)
  • N. Vadivelu et al.

    Peripheral opioid receptor agonists for analgesia: a comprehensive review

    J. Opioid Manag.

    (2011)
  • M.R. Bruchas et al.

    Stress-induced p38 mitogen-activated protein kinase activation mediates kappa-opioid-dependent dysphoria

    J. Neurosci.

    (2007)
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