Isotonitazene: A Novel Benzimidazole µ- Opioid Agonist as the Latest Novel Opioid
BK Logan1,2, AJ Krotulski2, DM Papsun1, SL Kacinko1 NMS Labs1 and CFSRE2 Disclosures
• Dr Logan is Executive Director of the Center for Forensic Science Research and Education, a 501 (3)(c) non-profit operating Foundation, which operates NPS Discovery ®. • He is an employee of NMS Labs, a commercial drug testing laboratory performing forensic and clinical toxicology and seized-drug testing, which provided data included in this presentation. Benzimidazole Analgesics
• US Patent 2,944,062, issued 1958 to Karl Hoffman, and Alfred Hunger at Ciba Pharmaceuticals, NJ. • Benzimidazoles identified as having good analgesic action. • Also claimed to have muscle relaxant properties. • Especially effective were those with nitro substitution at R2, and lower alkoxy group at R1 Benzimidazole Analgesics
• Etonitazene, clonitazene, isotonitazene all described in 1957 patent. • Highly potent benzimidazole µ- opioid agonists. etonitazene clonitazene • QSAR studies showed the significance of a particular A-F ring conformation in conferring high potency. • Etonitazene 60 times the potency of morphine in humans. isotonitazene
Feinberg AP, Creese I, Snyder SH. The opiate receptor: a model explaining structure-activity relationships of opiate agonists and antagonists. Proc Natl Acad Sci U S A. 1976;73(11):4215-4219. Benzimidazoles in Illicit Markets Benzimidazoles in Illicit Markets Benzimidazoles in Illicit Markets First Reported Benzimidazole Death
• Novel Psychoactive Substances (NPS) Detection in Alberta Casework (August 2019 update) • Brooks-Lim, Chatterton • Isotonitazene toxicity death in Deer Run, Alberta from March 2019 • Isotonitazene 0.12-0.56 ng/mL • Mesonitazene also positive in paraphernalia. Isotonitazene
• First synthesized and reported in the literature in the late 1950s. • Isotonitazene is a potent synthetic opioid related to synthetic opioids etonitazene, metonitazene, and clonitazene. • Pharmacological data suggest that this group of synthetic opioids have potency similar to or greater than fentanyl. • Isotonitazene estimated to have potency of 200-500X that of morphine in animal models. Isotonitazene Metabolism
N-Dealkylation
Nitro Reduction
O-Dealkylation N,O-DiDealkylation
• Metabolites M1, M2 and M3 frequently present in urine, not in blood • Hydrolysis had minimal impact on peak area
Krotulski AJ, Papsun DM, Kacinko SL, Logan BK. Isotonitazene Quantitation and Metabolite Discovery in Authentic Forensic Casework. J Anal Toxicol. 2020;44(6):521-530. Isotonitazene - Analysis
• Screening – NMS Labs • Confirmation - CFSRE • L/L extraction (n-BuCl/EtOAc) • L/L extraction (n-BuCl/EtOAc) • Analysis by LC-TOF-MS • Analysis by LC-MS-MS • Positive electrospray ionization • Fentanyl –D5 Istd • Mass range m/z 100 – 1000 • Linear Range 0.2-100ng/mL • Exact Mass (H+) 411.2391 Daltons • LOD – 0.02ng/mL • RT 5.656 mins • Standard Addition • LOD – 0.5ng/mL • Blank plus 3 spikes • Correlation R2>0.98
Krotulski AJ, Papsun DM, Kacinko SL, Logan BK. Isotonitazene Quantitation and Metabolite Discovery in Authentic Forensic Casework. J Anal Toxicol. 2020;44(6):521-530. Isotonitazene – Confirmation
Characteristic Evaluation Linearity (0.2-100 ng/mL) Linear (R2=0.999) Limit of Detection (LOD) <0.02 ng/mL Recovery 89% Matrix Interferences No interference Analyte Interferences No interference ISTD Interferences No interference
Krotulski AJ, Papsun DM, Kacinko SL, Logan BK. Isotonitazene Quantitation and Metabolite Discovery in Authentic Forensic Casework. J Anal Toxicol. 2020;44(6):521-530. Postmortem Case Findings
• N=150 positives, 116 quantitated • Blood concentration range: 0.19ng/mL – 39ng/mL • Mean/Median: 1.88+4.2ng/mL / 0.94ng/mL Postmortem Case Findings
• Most common combinations: • Flualprazolam (69%) • Etizolam (25%) • In ~half the cases, Isotonitazene was the only opioid • Cause of death was typically single or multiple drug intoxication • Manner of death was typically accidental Retrospective TOF Data-Mining
DATA-MINING SAMPLE-MINING PAST: Retrospective PRESENT: Real-Time Reanalysis
Sample Extracts
When was Increasing the The substance likelihood of first present in identifying the Analytical Data biological samples? substance close to the time of the case April 2019
16 July 2019
17 August 2019
18 September 2019
19 October 2019
20 November 2019
21 December 2019
22 January 2020
23 February 2020
24 March 2020
25 April 2020
26 May 2020
27 June 2020
28 July 2020
29 Total Isotonitazene cases by month
30 Isotonitazene to Brorphine Transition
31 Acknowledgements
• Alex Krotulski PhD, CFSRE • Donna Papsun MS, NMS Labs • Sherri Kacinko PhD, NMS Labs
• Judith Rodriquez –Salas MS, CFSRE [email protected] www.npsdiscovery.org