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. 2023 Oct 3;28(19):6925.
doi: 10.3390/molecules28196925.

Discovery of 7-Azanorbornane-Based Dual Agonists for the Delta and Kappa Opioid Receptors through an In Situ Screening Protocol

Fumika Karaki  1   2 , Taro Takamori  1 , Koumei Kawakami  1 , Sae Sakurai  1 , Kyoko Hidaka  1 , Kei Ishii  1 , Tomoya Oki  1 , Noriko Sato  3 , Nao Atsumi  1   4 , Karin Ashizawa  1   4 , Ai Taguchi  1   4 , Asuka Ura  1   4 , Toko Naruse  1 , Shigeto Hirayama  1   2 , Miki Nonaka  4 , Kanako Miyano  4 , Yasuhito Uezono  4 , Hideaki Fujii  1   2
Affiliations

Discovery of 7-Azanorbornane-Based Dual Agonists for the Delta and Kappa Opioid Receptors through an In Situ Screening Protocol

Fumika Karaki et al. Molecules. .

Abstract

In medicinal chemistry, the copper-catalyzed click reaction is used to prepare ligand candidates. This reaction is so clean that the bioactivities of the products can be determined without purification. Despite the advantages of this in situ screening protocol, the applicability of this method for transmembrane proteins has not been validated due to the incompatibility with copper catalysts. To address this point, we performed ligand screening for the µ, δ, and κ opioid receptors using this protocol. As we had previously reported the 7-azanorbornane skeleton as a privileged scaffold for the G protein-coupled receptors, we performed the click reactions between various 7-substituted 2-ethynyl-7-azanorbornanes and azides. Screening assays were performed without purification using the CellKeyTM system, and the putative hit compounds were re-synthesized and re-evaluated. Although the "hit" compounds for the µ and the δ receptors were totally inactive after purifications, three of the four "hits" for the κ receptor were true agonists for this receptor and also showed activities for the δ receptor. Although false positive/negative results exist as in other screening projects for soluble proteins, this in situ method is effective in identifying novel ligands for transmembrane proteins.

Keywords: 7-azanorbornane; G protein-coupled receptor; addiction; analgesics; click reaction; dual agonist; in situ screening; opioid receptor; polypharmacology.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Scheme 1
Scheme 1
Construction of the 7-azanorbornane-based compound library constructed by the click reactions. Structures of the KOR agonists identified in our previous study are also shown.
Figure 1
Figure 1
Schematic outline of library construction and first screening assay using the CellKeyTM system.
Figure 2
Figure 2
Interaction between MP1104, a KOR agonist with the morphinan scaffold (black), and the residues in KOR (red) [28]. The hydrogen-bonding water molecules are depicted in blue. The A-ring of MP1104 is indicated in the structure.
Figure 3
Figure 3
Agonistic activities of the triazoles and the parental alkynes and azides toward the opioid receptors. The positive controls are DAMGO (for MOR), SNC80 (for DOR), and (–)-U-50,488H (for KOR). All the compounds, including the positive controls were treated at the concentration of 10 µM.
Scheme 2
Scheme 2
Syntheses of the selected triazoles to validate the activities.
Figure 4
Figure 4
Agonistic activities in the presence and absence of selective antagonists for (a) DOR and (b) KOR. The concentrations of both the agonists and the antagonists were 10−5 M.
Figure 5
Figure 5
The dose-response curves of the triazoles and the alkynes for (a) DOR and (b) KOR.
Scheme 3
Scheme 3
Preparation of the alkynes to be used in the click reactions.
See this image and copyright information in PMC

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