Characterisation of AMB-FUBINACA metabolism and CB1-mediated activity of its acid metabolite

Hunter D J Webb; David B Finlay; Shuli Chen; Andrea J Vernall; Eric Sparkes; Samuel D Banister; Rhonda J Rosengren; Michelle Glass

doi:10.1007/s11419-022-00649-3

Characterisation of AMB-FUBINACA metabolism and CB₁-mediated activity of its acid metabolite

Forensic Toxicol. 2023 Jan;41(1):114-125. doi: 10.1007/s11419-022-00649-3. Epub 2022 Oct 28.

Authors

Hunter D J Webb¹, David B Finlay¹, Shuli Chen¹, Andrea J Vernall², Eric Sparkes^{3

4}, Samuel D Banister^{3

4}, Rhonda J Rosengren¹, Michelle Glass⁵

Affiliations

¹ Department of Pharmacology and Toxicology, University of Otago, Dunedin, 9016, New Zealand.
² Department of Chemistry, University of Otago, Dunedin, 9016, New Zealand.
³ Lambert Initiative for Cannabinoid Therapeutics, Brain and Mind Centre, The University of Sydney, Sydney, NSW, 2050, Australia.
⁴ School of Chemistry, The University of Sydney, Sydney, NSW, 2006, Australia.
⁵ Department of Pharmacology and Toxicology, University of Otago, Dunedin, 9016, New Zealand. michelle.glass@otago.ac.nz.

Abstract

Purpose: AMB-FUBINACA is a synthetic cannabinoid receptor agonist (SCRA) which is primarily metabolised by hepatic enzymes producing AMB-FUBINACA carboxylic acid. The metabolising enzymes associated with this biotransformation remain unknown. This study aimed to determine if AMB-FUBINACA metabolism could be reduced in the presence of carboxylesterase (CES) inhibitors and recreational drugs commonly consumed with it. The affinity and activity of the AMB-FUBINACA acid metabolite at the cannabinoid type-1 receptor (CB₁) was investigated to determine the activity of the metabolite.

Methods: The effect of CES1 and CES2 inhibitors, and delta-9-tetrahydrocannabinol (Δ⁹-THC) on AMB-FUBINACA metabolism were determined using both human liver microsomes (HLM) and recombinant carboxylesterases. Radioligand binding and cAMP assays comparing AMB-FUBINACA and AMB-FUBINACA acid were carried out in HEK293 cells expressing human CB₁.

Results: AMB-FUBINACA was rapidly metabolised by HLM in the presence and absence of NADPH. Additionally, CES1 and CES2 inhibitors both significantly reduced AMB-FUBINACA metabolism. Furthermore, digitonin (100 µM) significantly inhibited CES1-mediated metabolism of AMB-FUBINACA by ~ 56%, while the effects elicited by Δ⁹-THC were not statistically significant. AMB-FUBINACA acid produced only 26% radioligand displacement consistent with low affinity binding. In cAMP assays, the potency of AMB-FUBINACA was ~ 3000-fold greater at CB₁ as compared to the acid metabolite.

Conclusions: CES1A1 was identified as the main hepatic enzyme responsible for the metabolism of AMB-FUBINACA to its less potent carboxylic acid metabolite. This biotransformation was significantly inhibited by digitonin. Since other xenobiotics may also inhibit similar SCRA metabolic pathways, understanding these interactions may elucidate why some users experience high levels of harm following SCRA use.

Keywords: AMB-FUBINACA; Cannabinoid receptor; Carboxylesterase 1; Carboxylic acid metabolite; Human liver microsomes; Synthetic cannabinoid receptor agonist.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cannabinoid Receptor Agonists / pharmacology
Cannabinoids* / pharmacology
Digitonin
Dronabinol
HEK293 Cells
Humans

Substances

Cannabinoids
Dronabinol
methyl 2-(1-(4-fluorobenzyl)-1H-indazole-3-carboxamido)-3-methylbutanoate
Digitonin
Cannabinoid Receptor Agonists