Synthesis and in Vitro Cannabinoid Receptor 1 Activity of Recently Detected Synthetic Cannabinoids 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

Abstract

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.

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 II: Structure activity relationship assessment via a β‐arrestin recruitment assay

“…The EC 50 values for both reference compounds CP‐55,940 ( I ) (0.962 nM, 95% CI: 0.727–1.27) and JWH‐018 ( X ) (17.8 nM, 95% CI: 10.1–32.7) were in good agreement with previously reported data 3,13,14,19,29 . Very recently, Krotulski et al reported the pharmacological parameters for CB 1 for three of the tested substances, assessed using the same live cell‐based βarr2 recruitment assay, with results in good agreement with the present study: MMB‐4en‐PICA ( 1 ) (EC 50 : 125 nM, E max : 191% relative to 100% JWH‐018), MDMB‐4en‐PICA ( 2 ) (EC 50 : 11.5 nM, E max : 302% relative to 100% JWH‐018), and MDMB‐4en‐PINACA ( 12 ) (EC 50 : 2.47 nM, E max : 239% relative to 100% JWH‐018) 15,16 …”

“…Following activation of CB 1 , βarr2 is recruited to the activated receptor, resulting in functional complementation of NanoLuc, which, in the presence of a suitable substrate, will result in luminescence. This assay has extensively been used for the functional characterization of a wide panel (>100) of SCRAs, in a multitude of publications 3,13–16 . The outcome of this functional evaluation was compared to existing structure activity data from literature.…”

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 I—Synthesis, analytical characterization, and binding affinity for human CB1 receptors

“…), 110.5 (CH), 57.0 (CH), 52.3 (OCH 3 ), 46.2 (NCH 2 ), 31.9 (CH), 30.8 (CH 2 ), 29.0 (CH 2 ), 19.3 (CH 3 ), 18.2 (CH 3 ); v max 3331, 2962, 1739, 1629, 1508, 1466, 1388, 1279, 1229, 1175, 1146, 984, 911, 813, 780, 750, 636, 566, 541; HRMS (ESI+) C 20 H 26 N 2 O 3 [M + H] + exact mass 343.2022, accurate mass 343.2025 (mass error 2.49 ppm). All physical and spectral properties were consistent with previous reports 30 …”

“…), 30.9 (CH 2 ), 28.9 (CH 2 ), 26.8 (CH 3 ); v max 3413, 2962, 1729, 1700, 1520, 1490, 1367, 1329, 1214, 1166, 988, 910, 833, 775, 755, 555; HRMS (ESI+) C 20 H 27 N 3 O 3 [M + H] + exact mass 358.2131, accurate mass 358.2134 (mass error 2.51 ppm). All physical and spectral properties were consistent with previous reports 30 …”

Assessment of select synthetic cannabinoid receptor agonist bias and selectivity between the type 1 and type 2 cannabinoid receptor

“…As a limitation of our data, equivalent mouse pharmacokinetic data do not exist for 4-fluoro MDMB-BINACA and JWH 018 2′-naphthyl-N-(3-methylbutyl) and such data are beyond the scope of the present study. However, existing data presented here and elsewhere indicate that differences in SCRAs activity are pharmacodynamic rather than pharmacokinetic 45 – 47 . Although previous studies have not correlated the βarrestin2 bias of such ligands with specific in vivo effects, it is possible that βarrestin2-biased signaling may selectively enhance hypothermic and cataleptic responses relative to other cannabimimetic effects such as anti-nociception.…”