New synthetic cannabinoids carrying a cyclobutyl methyl side chain: Human Phase I metabolism and data on human cannabinoid receptor 1 binding and activation of Cumyl‐CBMICA and Cumyl‐CBMINACA

Abstract

Synthetic cannabinoids (SCs) represent a large group of new psychoactive substances (NPS), sustaining a high prevalence on the drug market since their first detection in 2008. Cumyl-CBMICA and Cumyl-CBMINACA, the first representatives of a new subclass of SCs characterized by a cyclobutyl methyl (CBM) moiety, were identified in July 2019 and February 2020. This work aimed at evaluating basic pharmacological characteristics and human phase-I-metabolism of these compounds. Human phase-I metabolites were tentatively identified by liquid-chromatographyquadrupole-time-of-flight-mass-spectrometry (LC-QToF-MS) of urine samples and confirmed by a pooled-human-liver-microsome (pHLM) assay. The basic pharmacological evaluation was performed applying a competitive ligand binding assay and a functional activation assay (GTPγS) using cell membranes carrying the human cannabinoid receptor 1 (hCB1). Investigation of the human phase-I-metabolism resulted in the identification of specific urinary markers built by mono-or dihydroxylation. While Cumyl-CBMICA was primarily hydroxylated at the indole ring, hydroxylation of Cumyl-CBMINACA mainly occurred at the CBM moiety. Both substances acted as agonists at the hCB1 receptor, although substantial differences could be observed. Cumyl-CBMINACA showed higher binding affinity (Ki = 1.32 nM vs. 29.3 nM), potency (EC50 = 55.4 nM vs. 497 nM) and efficacy (Emax = 207% vs. 168%) than its indole counterpart Cumyl-CBMICA.This study confirms that substitution of an indole by an indazole core tends to increase in vitro potency which is potentially reflected by higher in vivo potency. The emergence and disappearance of SCs distributed via online shops carrying a CBM moiety once more demonstrates the "cat-and-mouse" game between manufacturers and legislation.

Structure–activity relationships of valine, tert-leucine, and phenylalanine amino acid-derived synthetic cannabinoid receptor agonists related to ADB-BUTINACA, APP-BUTINACA, and ADB-P7AICA

“…In vitro binding data were acquired via a competitive radio binding assay, using radiolabelled [ 3 H]CP55,940 and human CB 1 and CB 2 receptor membrane preparations of HEK293 cells, as previously described. [33][34][35] The ability of the synthesised compounds to displace radioligands was measured and K i was determined (Table 1). The concentration displacement curves are shown in Fig.…”

The ADEBAR project: European and international provision of analytical data from structure elucidation and analytical characterization of NPS

“…The pharmacological parameters of receptor binding affinity and intrinsic activity of SCs are investigated within the ADEBAR project at the Institute of Forensic Medicine, Medical Center, University of Freiburg. 22 The investigated substances include well-known SCs as references, newly emerged SCs, 23,24 and specifically synthesized SC derivatives. A filtration-based radioligand assay is employed for both affinity and activity measurements.…”

Structure elucidation of the novel synthetic cannabinoid Cumyl‐Tosyl‐Indazole‐3‐Carboxamide (Cumyl‐TsINACA) found in illicit products in Germany

“…Especially with synthetic cannabimimetics (SC), as the largest and most diverse group of NPS, 1 the development of novel structures is driven by the pursuit of temporary legal substances distributed in pure form or as herbal blends, e‐liquids and impregnated paper 2–7 . In 2019, three SCs featuring a cyclobutyl methyl side chain were available to customers and detected in seizures by German forensic institutes 8,9 . Similarly, in 2020, the design of a norbornyl methyl side chain surfaced on the drug market in Germany, and the structures of three derivatives were elucidated 10 .…”

“…[2][3][4][5][6][7] In 2019, three SCs featuring a cyclobutyl methyl side chain were available to customers and detected in seizures by German forensic institutes. 8,9 Similarly, in 2020, the design of a norbornyl methyl side chain surfaced on the drug market in Germany, and the structures of three derivatives were elucidated. 10 Upon occurrence, these two side chain moieties were not scheduled under the German New Psychoactive Substance Act (NpSG) and the German Narcotics Act (BtMG).…”

“…This is not surprising given that pharmacological data of SCs show a trend towards greater potency at the hCB1 receptor for indazoles compared to indoles. 9,[11][12][13] already In 2020, 11 new SCs were notified, where indole core structures were still present next to indazoles.…”