Synthetic cannabinoid receptor agonists (SCRAs) are an evolving class of new psychoactive substances (NPS) with structurally diverse compounds emerging each year. Due to the rapid pace at which these drugs enter the market, there is often little or nil information regarding the pharmacology of these substances despite widespread human use. In this study, 12 recently emerged SCRAs (reported between 2018 and 2020) were synthesized, analytically characterized, and pharmacologically evaluated using a live cell-based nanoluciferase complementation reporter assay that monitors in vitro cannabinoid receptor type 1 (CB1) activation via its interaction with β-arrestin 2 (βarr2). All synthesized SCRAs acted as agonists of CB1, although differences in potency (EC50 = 2.33–5475 nM) and efficacy (E max = 37–378%) were noted, and several structure–activity relationships were identified. SCRAs featuring indazole cores (EC50 = 2.33–159 nM) were generally of equal or greater potency than indole analogues (EC50 = 32.9–330 nM) or 7-azaindole derivatives (EC50 = 64.0–5475 nM). Interestingly, with the exception of APP-BINACA (E max = 75.7%) and 5F-A-P7AICA (E max = 37.4%), all SCRAs showed greater efficacy than the historical SCRA JWH-018 to which responses were normalized (E max = 142–378%). The most potent CB1 agonists in the study were ADB-BINACA (EC50 = 6.36 nM), 4F-MDMB-BINACA (EC50 = 7.39 nM), and MDMB-4en-PINACA (EC50 = 2.33 nM). Notably, all of these SCRAs featured an indazole core as well as a “bulky” tert-butyl moiety in the pendant amino acid side chain. This study confirms that recently detected SCRAs 4F-MDMB-BICA, 5F-MPP-PICA, MMB-4en-PICA, CUMYL-CBMICA, ADB-BINACA, APP-BINACA, 4F-MDMB-BINACA, MDMB-4en-PINACA, A-CHMINACA, 5F-AB-P7AICA, 5F-MDMB-P7AICA, and 5F-AP7AICA were all able to activate the CB1 receptor in vitro, albeit to different extents, and are potentially psychoactive in vivo. These results indicate that further evaluation of these widely used NPS is warranted to better understand the risks associated with human consumption of these drugs.

Synthetic cannabinoid receptor agonists (SCRAs) are one of the largest and most structurally diverse classes of new psychoactive substances (NPS). Despite this, pharmacological data are often lacking following the identification of a new SCRA in drug markets. In this first of a three-part series, we describe the synthesis, analytical characterization, and binding affinity of a proactively generated, systematic library of 30 indole, indazole, and 7-azaindole SCRAs related to MMB-4en-PICA, MDMB-4en-PINACA, ADB-4en-PINACA, and MMB-4CN-BUTINACA featuring a 4-pentenyl (4en-P), butyl (B/BUT), or 4-cyanobutyl (4CN-B/BUT) tail and a methyl L-valinate (MMB), methyl L-tert-leucinate (MDMB), methyl L-phenylalaninate (MPP), L-valinamide (AB), L-tert-leucinamide (ADB), L-phenylalaninamide (APP), adamantyl (A), or cumyl head group. Competitive radioligand binding assays demonstrated that the indazole core conferred the highest CB 1 binding affinity (K i = 0.17-39 nM), followed by indole-(K i = 0.95-160 nM) and then 7-azaindole-derived SCRAs (K i = 5.4-271 nM). Variation of the head group had the greatest effect on binding, with tert-leucine amides and methyl esters (K i = 0.17-14 nM) generally showing the greatest affinities, followed by valine derivatives (K i = 0.72-180 nM), and then phenylalanine derivatives (K i = 2.5-271 nM). Adamantyl head groups (K i = 8.8-59 nM)

Synthetic cannabinoid receptor agonists (SCRAs) remain one the most prevalent classes of new psychoactive substances (NPS) worldwide, and examples are generally poorly characterised at the time of first detection. We...

Synthetic cannabinoid receptor agonists (SCRAs) are the second largest class of new psychoactive substances (NPS) and are associated with serious adverse effects and even death. Despite this, little pharmacological data are available for many of the most recent SCRAs. This study consists of three different parts, aiming to systematically evaluate a panel of 30 SCRAs using binding and different in vitro human cannabinoid 1 receptor (CB1) activation assays. The present Part II investigated the SCRA analogs for their CB1 activation via a β‐arrestin recruitment assay. The panel was systematically designed to include key structural sub‐features of recent SCRAs. Thus, the 4‐pentenyl tail of MMB‐4en‐PICA and MDMB‐4en‐PINACA was retained while incorporating varying head groups from other prevalent SCRAs, including amides and esters of L‐valine, L‐tert‐leucine, and L‐phenylalanine, and adamantyl and cumyl moieties. All 30 SCRAs activated CB1, with indazoles generally showing the greatest potency (EC50 = 1.88–281 nM), followed by indoles (EC50 = 11.5–2293 nM), and the corresponding 7‐azaindoles (EC50 = 62.4–9251 nM). Several subunit‐linked structure–activity relationships were identified: (i) tert‐leucine‐functionalized SCRAs were more potent than the corresponding valine derivatives; (ii) no major difference in potency or efficacy was observed between tert‐leucine/valine‐derived amides and the corresponding methyl esters; however, phenylalanine analogs were affected by this change; and (iii) minor structural changes to the 4‐pentenyl substituent had little influence on activity. These findings elucidate structural features that modulate the CB1 activation potential of currently prevalent SCRAs and a systematic panel of analogs, some of which may appear in NPS markets in future.

Synthetic cannabinoid receptor agonists (SCRAs) represent the most rapidly expanding class of new psychoactive substances (NPSs). Despite the prevalence and potency of recent chiral indole-3-carboxamide SCRAs, few pharmacological data are available regarding the enantiomeric bias of these NPSs toward human CB1 and CB2 receptors. A series of homochiral indole-3-carboxamides derived from (S)- and (R)-α-methylbenzylamine and featuring variation of the 1-alkyl substituent were prepared, pharmacologically evaluated, and compared to related achiral congeners derived from cumyl- and benzylamine. Competitive binding assays demonstrated that all analogues derived from either enantiomer of α-methylbenzylamine (14–17) showed affinities for CB1 (K i = 47.9–813 nM) and CB2 (K i = 47.9–347 nM) that were intermediate to that of the corresponding benzylic (10–13, CB1 K i = 550 nM to >10 μM; CB2 K i = 61.7 nM to >10 μM) and cumyl derivatives (6–9, CB1 K i = 12.6–21.4 nM; CB2 K i = 2.95–24.5 nM). In a fluorometric membrane potential assay, all α-methylbenzyl analogues (excluding 17) were potent, efficacious agonists of CB1 (EC50 = 32–464 nM; E max = 89–104%) and low efficacy agonists of CB2 (EC50 = 54–500 nM; E max = 52–77%), with comparable or greater potency than the benzyl analogues and much lower potency than the cumyl derivatives, consistent with binding trends. The relatively greater affinity and potency of (S)-14–17 compared to (R)-14–17 analogues at CB1 highlighted an enantiomeric bias for this series of SCRAs. Molecular dynamics simulations provided a conformational basis for the observed differences in agonist potency at CB1 pending benzylic substitution.

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 (CB 1 ) 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 Gα i protein (mini-Gα i assay), or binding of [ 35 S]-GTPγS. The observed efficacies (E max ) varied depending on the conducted assay. Statistical analysis suggests that the population means of the relative intrinsic activity (RA i ) significantly differ for the [ 35 S]-GTPγS assay and the other two assays, but the population means of the βarr2 and mini-Gα i assays were not statistically different. Our data suggest that differences observed between the βarr2 and mini-Gα i assays are the best The contribution to this article by Patrick Juchli is independent from and unrelated to his function at PwC Switzerland.

Synthetic cannabinoid receptor agonists (SCRAs) are a diverse class of new psychoactive substances (NPS). They commonly comprise N-alkylated indole, indazole, or 7-azaindole scaffolds with amide-linked pendant amino acid groups. To explore the contribution of the amino acid side chain to the cannabinoid pharmacology of SCRA NPS, a systematic library of side chainmodified SCRAs was prepared based on the recent detections of amino acid derivatives 17 (5F-AB-PINACA), 18 (5F-ADB-PINACA), 15 (PX-1), 19 (PX-2), and 20 (NNL-1). In vitro binding affinities and functional activities at cannabinoid type 1 and 2 receptors (CB 1 and CB 2 , respectively) were determined for all the library members using radioligand competition experiments and a fluorescence-based membrane potential assay. Binding affinities and functional activities varied widely across compounds (K i = 0.32 to >10 000 nM, EC 50 = 0.24−1259 nM), with several clear structure−activity relationships (SARs) emerging. Affinity and potency at CB 1 changed as a function of the heterocyclic core (indazole > indole > 7-azaindole) and the pendant amino acid side chain (tertbutyl > iso-propyl > iso-butyl > benzyl > ethyl > methyl > hydrogen). Ensemble docking at CB 1 revealed a clear steric basis for observed SAR trends. Interestingly, although 15 (PX-1) and 19 (PX-2) have been detected in recreational drug markets, they failed to induce centrally CB 1 -mediated effects (e.g., hypothermia) in mice using radiobiotelemetry. Together, these data provide insights regarding structural contributions to the cannabimimetic profiles of 17 (5F-AB-PINACA), 18 (5F-ADB-PINACA), 15 (PX-1), 19 (PX-2), 20 (NNL-1), and other SCRA NPS.

Synthetic cannabinoid receptor agonists (SCRAs) continue to make up a significant portion new psychoactive substances (NPS) detected and seized worldwide. Due to their often potent activation of central cannabinoid receptors in vivo, use of SCRAs can result in severe intoxication, in addition to other adverse health effects. Recent detections of AB-4CN-BUTICA, MMB-4CN-BUTINACA, MDMB-4F-BUTICA and MDMB-4F-BUTINACA mark a continuation in the appearance of SCRAs bearing novel tail substituents. The proactive characterization campaign described here has facilitated the detection of several new SCRAs in toxicological case work. Here we detail the synthesis, characterization, and pharmacological evaluation of recently detected SCRAs, as well as a systematic library of 32 compounds bearing head, tail, and core group combinations likely to appear in future. In vitro radioligand binding assays revealed most compounds showed moderate to high affinity at both CB1 (pKi = < 5 to 8.89 ± 0.09 M) and CB2 (pKi = 5.49 ± 0.03 to 9.92 ± 0.09 M) receptors. In vitro functional evaluation using a fluorescence-based membrane potential assay showed that most compounds were sub-micromolar to sub-nanomolar agonists at CB1 (pEC50 = < 5 to 9.48 ± 0.14 M) and CB2 (pEC50 = 5.92 ± 0.16 to 8.64 ± 0.15 M) receptors. An in silico receptor-ligand docking approach was utilized to rationalize binding trends for CB2 with respect to the tail substituent, and indicated that rigidity in this region (i.e., 4-cyanobutyl) was detrimental to affinity.