Phase I In Vitro Metabolic Profiling of the Synthetic Cannabinoid Receptor Agonists CUMYL-THPINACA and ADAMANTYL-THPINACA

Abstract

Synthetic cannabinoid receptor agonists (SCRAs) remain popular drugs of abuse. As many SCRAs are known to be mostly metabolized, in vitro phase I metabolic profiling was conducted of the two indazole-3-carboxamide SCRAs: CUMYL-THPINACA and ADAMANTYL-THPINACA. Both compounds were incubated using pooled human liver microsomes. The sample clean-up consisted of solid phase extraction, followed by analysis using liquid chromatography coupled to a high resolution mass spectrometer. In silico-assisted metabolite identification and structure elucidation with the data-mining software Compound Discoverer was applied. Overall, 28 metabolites were detected for CUMYL-THPINACA and 13 metabolites for ADAMATYL-THPINACA. Various mono-, di-, and tri-hydroxylated metabolites were detected. For each SCRA, an abundant and characteristic di-hydroxylated metabolite was identified as a possible in vivo biomarker for screening methods. Metabolizing cytochrome P450 isoenzymes were investigated via incubation of relevant recombinant liver enzymes. The involvement of mainly CYP3A4 and CYP3A5 in the metabolism of both substances were noted, and for CUMYL-THPINACA the additional involvement (to a lesser extent) of CYP2C8, CYP2C9, and CYP2C19 was observed. The results suggest that ADAMANTYL-THPINACA might be more prone to metabolic drug−drug interactions than CUMYL-THPINACA, when co-administrated with strong CYP3A4 inhibitors.

Defining Steric Requirements at CB1 and CB2 Cannabinoid Receptors Using Synthetic Cannabinoid Receptor Agonists 5F-AB-PINACA, 5F-ADB-PINACA, PX-1, PX-2, NNL-1, and Their Analogues

“…SCRAs featuring amino acid amides are subject to extensive metabolic biotransformation, ,, and it is possible that the low in vivo potency of 15 (PX-1) and 19 (PX-2) may be attributed to the rapid biotransformation of these compounds to inactive metabolites. Although 15 (PX-1) and 19 (PX-2) are closely related to several SCRAs that are potent CB 1 receptor agonists in vivo, even small changes in the molecular structure can produce vastly different metabolic profiles and clearance rates. − Species differences may also play a role in observed differences because in vitro potencies for the parent compounds at the human CB 1 receptor may not be reflected in mice given the highly idiosyncratic nature of structure–metabolism relationships for this class.…”

Synthesis and structure determination of a synthetic cannabinoid CUMYL‐THPINACA metabolite with differentiation between the ortho‐, meta‐, and para‐hydroxyl positions of the cumyl moiety

“…Conversely, the ortho and meta positions may be inaccessible to the enzyme due to steric hindrance, for example, nearby dimethyl groups. Monti et al also reported that only one metabolite of CUMYL‐THPINACA with mono‐hydroxylation at the cumyl moiety was detected 23 . However, several metabolites with mono‐hydroxylation at the cumyl moiety have been detected for another synthetic cannabinoids bearing the cumyl moiety 11–13 .…”

“…From another perspective, metabolite M3 was proposed to be 13 The ion detected at m/z 393.1997 was indicated as a structure in which one of the carbon atoms was present as a 13 C atom and a chemical formula of 13 mono-hydroxylation at the cumyl moiety was detected. 23 However, several metabolites with mono-hydroxylation at the cumyl moiety have been detected for another synthetic cannabinoids bearing the cumyl moiety. [11][12][13] The developed methods of synthesis and identification will be useful in identifying their structures through the differentiated position of the hydroxyl group at the cumyl moiety.…”

“…A previous study showed that one of the metabolites of CUMYL-THPINACA was mono-hydroxylated at the cumyl moiety. 23 In this study, metabolites with one hydroxyl group at the ortho, meta, or para position of the cumyl moiety were chemically synthesized to prepare the reference standards. The metabolite standards were analyzed via liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF/MS), which revealed differences in their retention time and product ion spectra.…”

“…CUMYL‐THPINACA is composed of a carboxamide‐type indazole core, tetrahydropyran‐4‐yl‐methyl (THPM) moiety, and cumyl moiety, which is the focus of this study. A previous study showed that one of the metabolites of CUMYL‐THPINACA was mono‐hydroxylated at the cumyl moiety 23 …”