Deltorphin, a novel amphibian skin peptide with high selectivity and affinity for δ opioid receptors

Kreil, Günther; Barra, Donatella; Simmaco, Maurizio; Erspamer, V.; Erspamer, G. Falconieri; Negri, Lucia; Severini, Cinzia; Corsi, Rita; Melchiorri, Pietro

doi:10.1016/0014-2999(89)90611-0

Cited by 224 publications

(95 citation statements)

References 5 publications

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“…Dermorphin, Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2, has high affinity and selectivity for the yu-opioid receptor (Broccardo et al, 1981;Yamashiro et al, 1983;Rossi et al, 1986;Amiche et al, 1990). In contrast, dermenkephalin, Tyr-D-Met-Phe-His-Leu-Met-Asp-NH2, and the deltorphins I and II, Tyr-D-Ala-Phe-aa-Val-Val-Gly-NH2 where 'aa' is either Asp or Glu, have high affinity and selectivity for the 6-opioid receptor Erspamer et al, 1989;Kreil et al, 1989;Lazarus et al, 1989); their affinities and selectivities are the equal of, or are superior to, those of the (Mosberg et al, 1983), (OtBu), Leu5]-enkephalyl-Thr6 and [DSer2, (OtBu), Leu5]-enkephalyl-Thr6 (Delay-Goyet et al, 1988).…”

Section: Introductionmentioning

confidence: 96%

“…A fourth group of opioid peptides has been found to be present in the skin of frogs belonging to the subfamily Phyllomedusinae Erspamer & Melchiorri, 1983;Erspamer et al, 1989). These heptapeptides, dermorphin , dermenkephalin Amiche et al, 1989) [also known as deltorphin or dermorphin gene-associated peptide (Lazarus et al, 1989)], deltorphin I and deltorphin II (Erspamer et al, 1989;Richter et al, 1990), are devoid of structural homology with mammalian opioids and contain a common N-terminal sequence Tyr-D-aa-Phe, in which 'D-aa' is either D-Ala or D-Met. Thus, they are unique among peptides synthesized by animal cells in having a D-amino acid in their sequence, even though the D-residue is encoded by the usual L-amino acid codons in the cDNA (Richter et al, 1987).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Opioid activity of dermenkephalin analogues in the guinea‐pig myenteric plexus and the hamster vas deferens

Sagan

Corbett

Amiche

et al. 1991

British J Pharmacology

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1 To elucidate the structural features required for selective and potent action of dermenkephalin at the 6-opioid receptor, a series of analogues of dermenkephalin and dermorphin were tested for their effectiveness in depressing electrically-evoked contractions of the vas deferens of the hamster (6-opioid receptors) and the guinea-pig myenteric plexus-longitudinal muscle preparation (u-and K-opioid receptors). 2 Dermenkephalin was more selective and more potent at 6-receptors than the 6-ligand [D-Pen2, DPen5]-enkephalin. The responses to dermenkephalin in the hamster vas deferens were increased by addition of peptidase inhibitors; the maximum effect was obtained with 3juM thiorphan.3 [L-Met2]-dermenkephalin had 0.2% and [L-Ala2]-dermorphin 0.01% of the agonist activity of the corresponding endogenous peptides which have D-amino acids in position 2. The pharmacological activity of these analogues was unaffected by inhibition of peptidases. This emphasizes the role that the Dconfiguration plays in determining the bioactive folding of these highly active peptides. 4 Dermenkephalin-(1-6)-NH2 was more potent at 6-receptors than at ,u-receptors whereas, dermenkephalin-(1-4)-NH2 is a selective u-agonist, having no activity at 6-receptors. 5 Substitution of the C-terminal tripeptide of dermorphin with the C-terminal tripeptide of dermenkephalin abolished the p-receptor preference of dermorphin. The resulting hybrid peptide, Tyr-D-Ala-PheGly-Leu-Met-Asp-NH2 was as potent as dermenkephalin at 6-receptors. A shift towards a preference for 6-receptors was obtained when the C-terminal tetrapeptide of dermorphin was replaced by the C-terminal tetrapeptide of dermenkephalin. 6 Substitution of Asp by Asn in position 7 of dermenkephalin caused an increase in p-receptor potency and a decrease in 6-receptor potency, resulting in a 20 fold decrease in p-receptor selectivity. Dermenkephalin-(1-6)-NH2 and [Asn7]-dermenkephalin have almost identical 6-receptor agonist potencies and ratios of IC50 in the myenteric plexus to IC50 in the hamster vas deferens. 7 The results obtained emphasise the importance of a negative charge at the C-terminus of dermenkephalin for selectivity at the 6-opioid receptor. Furthermore, the hydrophobic residues Leu5 and Met6 may be critical in ensuring tight binding to the receptor which results in high agonist potency.

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Section: Introductionmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

Opioid activity of dermenkephalin analogues in the guinea‐pig myenteric plexus and the hamster vas deferens

Sagan

Corbett

Amiche

et al. 1991

British J Pharmacology

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“…For comparison, a highly ␦-selective peptide, deltorphin-II (Tyr-D-Ala-Phe-Glu-Val-Val-Gly-NH 2 ), originating from amphibian skin (Kreil et al, 1989), has also been studied. We have used a functional assay based on agonist-evoked changes of intracellular Ca 2ϩ levels in CHO cells stably transfected with the opioid receptor ( or ␦) and apoaequorin cDNA.…”

Section: Discussionmentioning

confidence: 99%

Functional Characterization of Opioid Receptor Ligands by Aequorin Luminescence-Based Calcium Assay

Fichna¹,

Gach²,

Piestrzeniewicz³

et al. 2006

J Pharmacol Exp Ther

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A functional assay, based on aequorin-derived luminescence triggered by receptor-mediated changes in intracellular calcium levels, was used to examine relative potency and efficacy of the -opioid agonists endomorphin-1, endomorphin-2, morphiceptin, and their position 3-substituted analogs, as well as the ␦-agonist deltorphin-II. The results of the aequorin assay, performed on recombinant cell lines, were compared with those obtained in the functional assay on isolated tissue preparations (guinea pig ileum and mouse vas deferens). A range of nine opioid peptide ligands produced a similar rank order of potency for the -and ␦-opioid receptor agonists in both functional assays. The highest potency at the -receptor was observed for endomorphin-1, endomorphin-2, and [D-1-Nal 3 ]morphiceptin, whereas deltorphin-II was the most potent ␦-receptor agonist. In the aequorin assay, the -and ␦-agonist-triggered luminescence was inhibited by the opioid antagonists naloxone and naltrindole, respectively. We can conclude that the use of the aequorin assay for new -and ␦-receptor-selective opioid analogs gives pharmacologically relevant data and allows high-throughput compound screening, which does not involve radioactivity or animal tissues. This is the first study that validates the application of this assay in the screening of opioid analogs.Current methods for the identification and characterization of new ligands at the three main opioid receptor types (, ␦, ) typically use binding assays, where a radioligand with high affinity to one receptor type is displaced by a tested analog. Apart from binding studies, different functional assays can also be used to determine relative potency and efficacy of new analogs at the opioid receptors. The most popular functional assay used for opioid activity determination in vitro is performed on isolated tissue preparations. This assay is based on inhibition of electrically evoked contractions of guinea pig ileum (GPI) and mouse vas deferens (MVD). The opioid effect in the GPI preparations is mainly mediated by the -receptors, whereas the predominant receptors of the MVD are of the ␦-type. The GPI/MVD assay makes it possible to determine the -and ␦-receptor interactions, which is the main focus in standard tests for opioid activity.In the present study, we used an alternative functional assay, which does not involve radioactivity or animal tissues and allows high-throughput screening of opioid peptide analogs. This assay is based on the recombinant mammalian cell line expressing an opioid receptor together with a luminescent reporter protein.Opioid receptors belong to a large family of the G proteincoupled receptors (GPCR), which on stimulation by a ligand induce the activation of the phospholipase C, with the subsequent generation of the primary second messenger metabolites: diacylglycerol and inositol 1,4,5-trisphosphate, followed by a release of calcium from intracellular stores. These agonist-induced receptor-mediated changes in calcium levels can be quantitatively assessed and ...

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“…To validate the heteromer assay for this purpose, we tested four DOR-selective compounds for their activity on DOR homomers (DOR 333 -G qi4 1 DOR), MOR homomers (MOR 354 -G qi4 1 MOR), and DOR-MOR heteromers (DOR 333 -G qi4 1 MOR; MOR 354 -G qi4 1 DOR). The set included deltorphin II, an amphibian-derived peptide (Kreil et al, 1989); a DOR agonist currently in phase 2 clinical trials, ADL5859 (Le Bourdonnec et al, 2008); as well as two agonists, SNC80 and ARM1000390, that differ in their ability to internalize the DOR (Pradhan et al, 2009). Deltorphin II displayed similar activity on the DOR homomer and the DOR-MOR heteromers ( Fig.…”

Section: Screening For Homomer-and Heteromer-selective Compoundsmentioning

confidence: 99%

Novel Screening Assay for the Selective Detection of G-Protein-Coupled Receptor Heteromer Signaling

Rijn

Harvey

Brissett

et al. 2012

J Pharmacol Exp Ther

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Drugs targeting G-protein-coupled receptors (GPCRs) make up more than 25% of all prescribed medicines. The ability of GPCRs to form heteromers with unique signaling properties suggests an entirely new and unexplored pool of drug targets. However, current in vitro assays are ill equipped to detect heteromer-selective compounds. We have successfully adapted an approach, using fusion proteins of GPCRs and chimeric G proteins, to create an in vitro screening assay (in human embryonic kidney cells) in which only activated heteromers are detectable. Here we show that this assay can demonstrate heteromer-selective G-protein bias as well as measure transinhibition. Using this assay, we reveal that the d-opioid receptor agonist ADL5859, which is currently in clinical trials, has a 10-fold higher potency against d-opioid receptor homomers than d/m-opioid receptor heteromers (pEC 50 5 6.7 6 0.1 versus 5.8 6 0.2). The assay enables the screening of large compound libraries to identify heteromer-selective compounds that could then be used in vivo to determine the functional role of heteromers and develop potential therapeutic agents.

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Deltorphin, a novel amphibian skin peptide with high selectivity and affinity for δ opioid receptors

Cited by 224 publications

References 5 publications

Opioid activity of dermenkephalin analogues in the guinea‐pig myenteric plexus and the hamster vas deferens

Opioid activity of dermenkephalin analogues in the guinea‐pig myenteric plexus and the hamster vas deferens

Functional Characterization of Opioid Receptor Ligands by Aequorin Luminescence-Based Calcium Assay

Novel Screening Assay for the Selective Detection of G-Protein-Coupled Receptor Heteromer Signaling

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