Enzyme-Linked Immunosorbent Assay (ELISA) for the Detection of Use of the Synthetic Cannabinoid Agonists UR-144 and XLR-11 in Human Urine

Abstract

Ongoing changes in the synthetic cannabinoid drug market create the need for relevant targeted immunoassays for rapid screening of biological samples. We describe the validation and performance characteristics of an enzyme-linked immunosorbent assay designed to detect use of one of the most prevalent synthetic cannabinoids in urine, UR-144, by targeting its pentanoic acid metabolite. Fluorinated UR-144 (XLR-11) has been demonstrated to metabolize to this common product. The assay has significant cross-reactivity with UR-144-5-OH, UR-144-4-OH and XLR-11-4-OH metabolites, but <10% cross-reactivity with the parent compounds, and no measurable cross-reactivity with other synthetic cannabinoids and their metabolites at concentrations of <1,000 ng/mL. The assay's cutoff is 5 ng/mL relative to the pentanoic acid metabolite of UR-144, which is used as the calibrator. The method was validated with 90 positive and negative control urine samples for UR-144, XLR-11 and its metabolites tested versus liquid chromatography-tandem mass spectrometry. The accuracy, sensitivity and specificity were determined to be 100% for the assay at the specified cutoff.

Genotoxic properties of XLR-11, a widely consumed synthetic cannabinoid, and of the benzoyl indole RCS-4

“…These findings were not unexpected; also with other indole SCs including UR-144, which is structurally related to XLR-11 (XLR-11 is the fluorinated form of UR-144) (Mohr et al 2014), consistently negative results were obtained in earlier bacterial mutagenicity tests (Koller et al 2015); also the natural cannabinoid Δ 9 -THC does not cause induction of his + revertants in Salmonella tester strains (NTP 1996). SCs with benzoyl structure like RCS-4 have not been studied before in microbial mutagenicity assays according to our knowledge, and the present findings show that the compound is devoid of activity under all experimental conditions.…”

Δ9-Tetrahydrocannabinol-like effects of novel synthetic cannabinoids found on the gray market

“…Four of the six compounds (UR-144, XLR-11, AKB-48, and AB-FUBINACA) have been identified in samples of synthetic cannabinoids obtained on the street (Kavanagh et al, 2013; Uchiyama et al, 2013; Strano Rossi et al, 2014). All six of the compounds have been found in blood or urine samples (or identified by verbal report) of users reporting adverse effects (Behonick et al, 2014; Gugelmann et al, 2014; Mohr et al, 2014; Strano Rossi et al, 2014), and the use of UR-144 or XLR-11 has been reported in cases of driving under the influence (Lemos, 2014; Musshoff et al, 2014). Of further concern, several of these compounds have been reported to produce significant adverse effects.…”

Translational Metabolomics: Current Challenges and Future Opportunities

“…Waters has commercialised an ACQUITY UPLC I-Class/Xevo Tandem Quadrupole Detector (TQD) IVD System that provides robust analysis of metabolites in clinical laboratories (). Other simpler ways of quantifying smaller and targeted metabolites are the available Enzyme-linked immunosorbent assay (ELISA) and other enzymatic kits [141,142,143,144]. However, these approaches also have limitations including metabolite degradation, rapid metabolite turnover rates during the analysis, cross-reactivity, lack of simultaneous analysis and poor enzyme/antibody specificity [144,145].…”