Differential signaling properties at the kappa opioid receptor of 12-epi-salvinorin A and its analogues

Abstract

The kappa opioid receptor (KOPR) has been identified as a potential drug target to prevent or alter the course of mood, anxiety and addictive disorders or reduce response to stress. In a search for highly potent and selective KOPR partial agonists as pharmacological tools, we have modified 12-epi-salvinorin A, a compound which we have previously observed to be a KOPR partial agonist. Five analogues of 12-epi-salvinorin A were synthesized and their effects on G protein activation as well as β-arrestin2 recruitment were evaluated. Only 12-epi-salvinorin A (1) partially activated signaling through G proteins, yet acted as a full agonist in the β-arrestin 2 DiscoveRx assay. Other salvinorin analogues tested in these functional assays were full agonists in both assays of KOPR activation. By comparison, the non-selective opioid ligand nalbuphine, known to be a partial agonist for G-protein activation, was also a partial agonist for the β-arrestin mediated signaling pathway activated through KOPR.

Figure 1

Agonist-induced recruitment of βarr2-GFP to KOPR in U2OS cells. Live cell confocal imaging of KOPR-βarr2eGFP-U2OS cells reveals robust recruitment of β-arrestin 2 to the KOPR by salvA and its derivatives as evidenced by the formation of intense green punctae after 10 min of drug exposure. A 5× magnification of the region outlined by the white square in each image is shown on the right. SalvA and the two derivatives (1 and 3) (10 μM) robustly recruit β-arrestin 2 to the KOPR (bright punctae), while nalbuphine (10 μM), a partial agonist, recruits less βarr2-GFP. Similar results obtained from three independent experiments performed on separate days, representative images shown.

Scheme 1

(a) NaHCO₃, H₂O₂, THF, 58%; (b) MOMCl, NaI, DIPEA, DMA, 80 °C, 60%; (c) EtOMeCl, NaI, DIPEA, DMA, 80 °C, 65%; (d) NBS (1.1 equiv), CHCl₃, 28%; (e) NBS (2.2 equiv), CHCl₃, 31%.