Pharmacological Investigations of N-Substituent Variation in Morphine and Oxymorphone: Opioid Receptor Binding, Signaling and Antinociceptive Activity

Morphine and structurally related derivatives are highly effective analgesics, and the mainstay in the medical management of moderate to severe pain. Pharmacological actions of opioid analgesics are primarily mediated through agonism at the mopioid peptide (MOP) receptor, a G protein-coupled receptor. Position 17 in morphine has been one of the most manipulated sites on the scaffold and intensive research has focused on replacements of the 17-methyl group with other substituents. Structural variations at the N-17 of the morphinan skeleton led to a diversity of molecules appraised as valuable and potential therapeutics and important research probes. Discovery of therapeutically useful morphine-like drugs has also targeted the C-6 hydroxyl group, with oxymorphone as one of the clinically relevant opioid analgesics, where a carbonyl instead of a hydroxyl group is present at position 6. Herein, we describe the effect of N-substituent variation in morphine and oxymorphone on in vitro and in vivo biological properties and the emerging structure-activity relationships. We show that the presence of a N-phenethyl group in position 17 is highly favorable in terms of improved affinity and selectivity at the MOP receptor, potent agonism and antinociceptive efficacy. The N-phenethyl derivatives of morphine and oxymorphone were very potent in stimulating G protein coupling and intracellular calcium release through the MOP receptor. In vivo, they were highly effective against acute thermal nociception in mice with marked increased antinociceptive potency compared to the lead molecules. It was also demonstrated that a carbonyl group at position 6 is preferable to a hydroxyl function in these N-phenethyl derivatives, enhancing MOP receptor affinity and agonist potency in vitro and in vivo. These results expand the understanding of the impact of different moieties at the morphinan nitrogen on ligand-receptor interaction, molecular mode of action and signaling, and may be instrumental to the development of new opioid therapeutics

N-Phenethyl Substitution in 14-Methoxy-N-methylmorphinan-6-ones Turns Selective mu Opioid Receptor Ligands into Dual mu/delta Opioid Receptor Agonists

Morphine and structurally-derived compounds are mu opioid receptor (mu OR) agonists, and the most effective analgesic drugs. However, their usefulness is limited by serious side effects, including dependence and abuse potential. The N-substituent in morphinans plays an important role in opioid activities in vitro and in vivo. This study presents the synthesis and pharmacological evaluation of new N-phenethyl substituted 14-O-methylmorphinan-6-ones. Whereas substitution of the N-methyl substituent in morphine (1) and oxymorphone (2) by an N-phenethyl group enhances binding affinity, selectivity and agonist potency at the mu OR of 1a and 2a, the N-phenethyl substitution in 14-methoxy-N-methylmorphinan-6-ones (3 and 4) converts selective mu OR ligands into dual mu/delta OR agonists (3a and 4a). Contrary to N-methylmorphinans 1-4, the N-phenethyl substituted morphinans 1a-4a produce effective and potent antinociception without motor impairment in mice. Using docking and molecular dynamics simulations with the mu OR, we establish that N-methylmorphinans 1-4 and their N-phenethyl counterparts 1a-4a share several essential receptor-ligand interactions, but also interaction pattern differences related to specific structural features, thus providing a structural basis for their pharmacological profiles. The emerged structure-activity relationships in this class of morphinans provide important information for tuning in vitro and in vivo opioid activities towards discovery of effective and safer analgesics

Exploring pharmacological activities and signaling of morphinans substituted in position 6 as potent agonists interacting with the μ opioid receptor

Background: Opioid analgesics are the most effective drugs for the treatment of moderate to severe pain. However, they also produce several adverse effects that can complicate pain management. The μ opioid (MOP)receptor, a G protein-coupled receptor, is recognized as the opioid receptor type which primarily mediates the pharmacological actions of clinically used opioid agonists. The morphinan class of analgesics including morphine and oxycodone are of main importance as therapeutically valuable drugs. Though the natural alkaloid morphine contains a C-6-hydroxyl group and the semisynthetic derivative oxycodone has a 6-carbonyl function, chemical approaches have uncovered that functionalizing position 6 gives rise to a range of diverse activities. Hence, position 6 of N-methylmorphinans is one of the most manipulated sites, and is established to play a key role in ligand binding at the MOP receptor, efficacy, signaling, and analgesic potency. We have earlier reported on a chemically innovative modification in oxycodone resulting in novel morphinans with 6-acrylonitrile incorporated substructures. Results: This study describes in vitro and in vivo pharmacological activities and signaling of new morphinans substituted in position 6 with acrylonitrile and amido functions as potent agonists and antinociceptive agents interacting with MOP receptors. We show that the presence of a 6-cyano group in N-methylmorphinans has a strong influence on the binding to the opioid receptors and post-receptor signaling. One 6-cyano-N-methylmorphinan of the series was identified as the highest affinity and most selective MOP agonist, and very potent in stimulating G protein coupling and intracellular calcium release through the MOP receptor. In vivo, this MOP agonist showed to be greatly effective against thermal and chemical nociception in mice with marked increased antinociceptive potency than the lead molecule oxycodone. Conclusion: Development of such novel chemotypes by targeting position 6 provides valuable insights on ligand-receptor interaction and molecular mode of action, and may aid in identification of opioid therapeutics with enhanced analgesic properties and fewer undesirable effects

<i>In vitro</i> agonist potency and efficacy.

a<p>Membranes from CHO cells stably transfected with human MOP, DOP or KOP receptors were used.</p>b<p>CHO cells co-expressing chimeric G proteins and recombinant human MOP, DOP or KOP receptors.</p>c<p>E_max is expressed in percentage relative to maximal stimulation produced by DAMGO (MOP), DPDPE (DOP) or U69,593 (KOP).</p>d<p>Inactive up to 10 µM.</p>e<p>crc, concentration response curve.</p>f<p>ND, not determined due to very low binding affinity at the KOP receptor.</p><p>Values represent the mean ± SEM of at least three experiments each performed in duplicate or triplicate.</p

Dose-dependent antinociceptive effects produced by morphine, oxymorphone and <i>N</i>-methylmorphinans 1, 4 and 6.

<p>(A) Hot-plate test. (B) Tail-flick test. Hot-plate and tail-flick latencies (as %MPE) are shown at 30 min (peak of action) after s.c. drug administration to mice. Data are shown as the mean ± SEM (<i>n</i> = 5–6 mice per group).</p

Highly Potent and Selective New Diphenethylamines Interacting with the κ‑Opioid Receptor: Synthesis, Pharmacology, and Structure–Activity Relationships

We previously reported on a series of small molecules targeting the κ-opioid (KOP) receptor featuring a diphenethylamine scaffold and showed the promise of these ligands as effective analgesics with reduced liability for adverse effects. This study expands the structure–activity relationships on our original series by presenting several modifications in the lead compounds <b>1</b> (HS665) and <b>2</b> (HS666). A library of new diphenethylamines was designed, synthesized, and pharmacologically evaluated. In comparison with <b>1</b> and <b>2</b>, the KOP receptor affinity, selectivity, and agonist activity were modulated by introducing bulkier N-substituents, a 2-fluoro substitution, and additional hydroxyl groups at positions 3′ and 4′. Several analogues showed subnanomolar affinity and excellent KOP receptor selectivity acting as full or partial agonists, and one as an antagonist. The new diphenethylamines displayed antinociceptive efficacies with increased potencies than U50,488, <b>1</b> and <b>2</b> in the writhing assay and without inducing motor dysfunction after sc administration in mice

Structures of morphine, oxymorphone and N-substituted morphinans 1–6.

<p>Ph, phenyl.</p

Opioid receptor binding affinities and selectivities at MOP, DOP and KOP receptors.

<p>Binding assays were performed with membranes from rat brain (MOP and DOP receptors) and guinea pig brain (KOP receptors).</p><p>Values represent the mean ± SEM of at least three experiments each performed in duplicate.</p

Time-course of antinociceptive effects produced by morphine, oxymorphone and <i>N</i>-methylmorphinans 1, 4 and 6.

<p>The effect of morphine (1.25–5 mg/kg), oxymorphone (0.2–1 mg/kg), and compounds <b>1</b> (0.05–0.5 mg/kg), <b>4</b> (0.5–5 mg/kg), and <b>6</b> (0.1–0.5 mg/kg) in the hot-plate test (A, left panel) and in the tail-flick test (B, right panel). Hot-plate and tail-flick latencies (in seconds) were determined in mice before (0 min) and after s.c. drug administration (30, 60 and 120 min). Data are shown as the mean ± SEM (<i>n</i> = 5–6 mice per group).</p