Pharmacological characterization of the bifunctional opioid ligand H-Dmt-Tic-Gly-NH-Bzl (UFP-505) †

Abstract

We have shown in a range of preparations and assays that UFP-505 behaves as a potent MOP agonist and DOP antagonist; a MOP/DOP bifunctional opioid. Further studies in dual expression systems and whole animals with this prototype are warranted.

In vitro and in vivo characterization of the bifunctional μ and δ opioid receptor ligand UFP‐505

“…UFP‐505 induced a significant internalization of μ receptors at 10 and 1 μΜ compared to the control (untreated), with a pEC 50 of 6.62. This is similar with the pEC 50 produced from GTPγ 35 S binding (6.37) but lower than the pK i of UFP‐505 binding (7.79) as shown in previously published data (Dietis et al, ).…”

“…In part of the present study, we have used δ receptor‐expressing cells with receptor density (Figure A left ~1.8 pmol mg·protein −1 ), ~1.8 higher than the reported value in Dietis et al . (). In membranes prepared from these high‐δ receptor expressing cells, we found significant but low partial agonist activity for UFP‐505 in a GTPγ 35 S assay (α 0.28 relative to DPDPE), as shown in Figure A right, indicating a partial agonist activity at δ receptors, at very high (non‐physiological) receptor density.…”

“…CHO cells stably expressing a single type of human opioid receptor (μ receptors, hMOP and δ receptors, hDOP) and cells with high expression of human δ receptors (CHO hDOP/high ) were grown as described previously (Dietis et al, ). Stock cultures were supplemented with 200 μg·mL −1 geneticin (G418), for CHO hMOP , CHO hDOP and CHO hDOP/high cells, and with additional 200 μg·mL −1 hygromycin B for the novel CHO hMOP/hDOP cell line (see below).…”

“…Adherent cells were then washed three times at 4°C with harvest buffer (containing 0.9% saline, 0.02% EDTA, 10 mM HEPES, pH of 7.4) to remove any receptor‐bound ligand, before harvesting. Membranes were prepared as previously described (Dietis et al, ), washing three times with buffer prior to three centrifugations at 1600× g and three centrifugations at 14 000× g , with subsequent removal of supernatant and resuspension of the pellet, in order to ensure that all desensitizing ligand is washed off. Then a saturation binding assay was performed as described above.…”

“…Our group has used the bifunctional opioid H‐Dmt‐Tic‐Gly‐NH‐Bzl (UFP‐505) as a prototype μ receptor agonist and δ receptor antagonist (Balboni et al, ; Dietis et al, ) and hypothesized that this opioid ligand will produce antinociceptive actions in vivo with reduced analgesic tolerance liability. Here, we present our data from relevant in vivo assays on the tolerance, as well as new data on this ligand's effects on opioid receptor density and trafficking from assays in vitro, at recombinant μ and δ receptors and at μ /δ double expression receptor systems, and ex vivo , using rat tissues.…”