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Systematic evaluation of a panel of 30 synthetic cannabinoid receptor agonists structurally related to MMB-4en-PICA,MDMB-4en-PINACA,ADB-4en-PINACA,and MMB-4CN-BUTINACA using a combination of binding and different CB1 receptor activation assays. Part III: The G protein pathway and critical comparison of different assays |
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| Authors: | Katharina Elisabeth Grafinger Marthe M. Vandeputte Annelies Cannaert Adam Ametovski Eric Sparkes Elizabeth Cairns Patrick Osamu Juchli Belal Haschimi Benedikt Pulver Samuel D. Banister Christophe P. Stove Volker Auwärter |
| Affiliation: | 1. Institute of Forensic Medicine, Forensic Toxicology, Medical Center – University of Freiburg, Faculty of Medicine, University of Freiburg, Freiburg, Germany;2. Laboratory of Toxicology, Department of Bioanalysis, Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium;3. The Lambert Initiative for Cannabinoid Therapeutics, Brain and Mind Centre, The University of Sydney, Camperdown, NSW, Australia School of Chemistry, The University of Sydney, Sydney, NSW, Australia;4. The Lambert Initiative for Cannabinoid Therapeutics, Brain and Mind Centre, The University of Sydney, Camperdown, NSW, Australia;5. Data & Analytics, PwC Switzerland, Zürich, Switzerland |
| Abstract: | The present work is the last of a three-part study investigating a panel of 30 systematically designed synthetic cannabinoid receptor agonists (SCRAs) including features such as the 4-pentenyl tail and varying head groups including amides and esters of l -valine (MMB, AB), l -tert-leucine (ADB), and l -phenylalanine (APP), as well as adamantyl (A) and cumyl moieties (CUMYL). Here, we evaluated these SCRAs for their capacity to activate the human cannabinoid receptor 1 (CB1) via indirect measurement of G protein recruitment. Furthermore, we comparatively evaluated the results obtained from three in vitro assays, based on the recruitment of β-arrestin 2 (βarr2 assay) or Gαi protein (mini-Gαi assay), or binding of [35S]-GTPγS. The observed efficacies (Emax) varied depending on the conducted assay. Statistical analysis suggests that the population means of the relative intrinsic activity (RAi) significantly differ for the [35S]-GTPγS assay and the other two assays, but the population means of the βarr2 and mini-Gαi assays were not statistically different. Our data suggest that differences observed between the βarr2 and mini-Gαi assays are the best predictor for ‘biased agonism’ towards βarr or G protein recruitment in our study. SCRAs carrying an ADB or MPP moiety as a head group tended to produce elevated Emax values in the βarr2 assay, which might result in a tendency of these compounds to cause pronounced tolerance in users—a hypothesis that should be evaluated further by future studies. In general, a comparison of efficacies derived from different assays is difficult and should only be conducted very cautiously. |
| Keywords: | [35S]-GTPγS βarr2 recruitment Gαi recruitment structure activity relationship synthetic cannabinoid receptor agonists |