In vitro and in vivo human metabolism of a new synthetic cannabinoid NM-2201 (CBL-2201)

Abstract

In 2014, NM-2201 (CBL-2201), a novel synthetic cannabinoid (SC), was detected by Russian and United States laboratories. It was already added to the scheduled drugs list in Japan, Sweden and Germany. Unfortunately, no human metabolism data are currently available, making it challenging to confirm its intake because all previous investigated SCs were extensively metabolized. The present study aims to recommend appropriate marker metabolites by investigating NM-2201 metabolism in human hepatocytes and confirm the results in authentic human urine specimens. For the metabolic stability assay, 1 μM NM-2201 was incubated in human liver microsomes (HLMs) for up to 1 h; for metabolite profiling, 10 μM of NM-2201 was incubated in human hepatocytes for 3 h. Two authentic urine specimens from NM-2201 positive cases were analyzed after β-glucuronidase hydrolysis. Metabolite identification in hepatocyte samples and urine specimens was achieved with high-resolution mass spectrometry via information-dependent acquisition. NM-2201 was quickly metabolized in HLMs with an 8.0 min half-life. In human hepatocyte incubation samples, a total of thirteen NM-2201 metabolites were identified, generated mainly from ester hydrolysis and further hydroxylation, oxidative defluorination and subsequent glucuronidation. M13 (5-fluoro PB-22 3-carboxyindole) was the major metabolite. In the urine specimens, the parent drug NM-2201 was not detected; M13 was the predominant metabolite after β-glucuronidase hydrolysis. Therefore, based on our study, we recommend the M13 as a suitable urinary marker metabolite for confirming NM-2201 and/or 5F-PB-22 intake.

Identification of New Synthetic Cannabinoid ADB-CHMINACA (MAB-CHMINACA) Metabolites in Human Hepatocytes

“…In previous experiments involving human metabolism, hepatocyte incubations appeared successful to predict in vivo urinary metabolites (29)(30)(31)(32)42,43). Ideally, a confirmation of our results with urine specimens from authentic cases of ADB-CHMINACA intake would be advantageous (33); however, despite our efforts, we were unable to obtain such specimens.…”

In vitro metabolism of the synthetic cannabinoids PX‐1, PX‐2, and PX‐3 by high‐resolution mass spectrometry and their clearance rates in human liver microsomes

“…The half‐life of the control, naphthalen‐1‐yl 1‐(5‐fluoropentyl)‐1 H ‐indole‐3‐carboxylate (NM2201), was 6.7 ± 0.75 min and intrinsic clearance, 0.103 mL/min/mg. Diao et al reported a half‐life value of 8.0 ± 1.5 min and in vitro clearance as 0.088 mL/min/mg for NM2201 . The calculated half‐lives for PX‐1, PX‐2, and PX‐3 were 15.1 ± 1.02, 3.4 ± 0.27, and 5.2 ± 0.89, respectively, as shown in Table .…”

“…The reaction mixture contained 780 μL distilled water, 100 μL 0.5 M potassium phosphate buffer, pH 7.4, 10 μL solution B, 10 μL 100.0 μmol/L drug, 50 μL HLM suspension, and 50 μL solution A to initiate the reaction. NM2201 was used as a positive control and the metabolite 1‐(5‐fluoropentyl)‐1 H ‐indole‐3‐carboxylic acid (5FPB22) was monitored as previously studied . All samples were incubated at 37°C.…”

Approaches, Challenges, and Advances in Metabolism of New Synthetic Cannabinoids and Identification of Optimal Urinary Marker Metabolites

“…Human hepatocyte incubation and high‐resolution MS were utilized to reveal metabolites of multiple new SCs . The model successfully predicted urinary metabolites that were confirmed in urine specimens from suspected SC ingestion for FDU‐PB‐22 ( Figure a ), FUB‐PB‐22 ( Figure a ), NM‐2201 ( Figure b ), 5F‐AB‐PINACA ( Figure c ), AB‐FUBINACA ( Figure a ), and AB‐PINACA ( Figure b ) …”

“…For NM‐2201, the parent compound also was absent ( Figure b ) in the 3‐h hepatocyte incubation sample . NM‐2201 was extensively hydrolyzed via ester hydrolysis, generating the main metabolite M13 (5F‐PI‐COOH).…”