Pharmacological evaluation and forensic case series of N-pyrrolidino etonitazene (etonitazepyne), a newly emerging 2-benzylbenzimidazole ‘nitazene’ synthetic opioid

Abstract

Novel synthetic opioids continue to emerge on recreational drug markets worldwide. In response to legislative bans on fentanyl analogues, non-fentanyl structural templates, such as 2-benzylbenzimidazoles ('nitazenes'), are being exploited to create new -opioid receptor (MOR) agonists. Here, we pharmacologically characterize an emerging cyclic analogue of etonitazene, called N-pyrrolidino etonitazene (etonitazepyne), using in vitro and in vivo methods. A series of analytically confirmed fatalities is described to complement preclinical findings. Radioligand binding assays in rat brain tissue revealed that N-pyrrolidino etonitazene has high affinity for MOR (Ki=4.09 nM) over -opioid (Ki=959 nM) and -opioid (Ki=980 nM) receptors. In a MOR--arrestin2 activation assay, N-pyrrolidino etonitazene displayed high potency (EC50=0.348 nM), similar to etonitazene (EC50=0.360 nM), and largely exceeding the potencies of fentanyl (EC50=14.9 nM) and morphine (EC50=290 nM). When administered s.c. to male Sprague Dawley rats, N-pyrrolidino etonitazene induced opioid-like antinociceptive, cataleptic, and thermic effects. Its potency in the hot plate test (ED50=0.0017 mg/kg) was 10fold and 2,000-fold greater than fentanyl (ED50=0.0209 mg/kg) and morphine (ED50=3.940 mg/kg), respectively. Twenty-one overdose fatalities associated with N-pyrrolidino etonitazene were found to contain low blood concentrations of the drug (median=2.2 ng/mL), commonly in the context of polysubstance use. N-Pyrrolidino etonitazene was reported as a cause of death in at least two cases, demonstrating toxicity in humans. We demonstrate that N-pyrrolidino etonitazene is an extremely potent MOR agonist that is likely to present high risk to users. Continued vigilance is required to identify and characterize emergent 2benzylbenzimidazoles, and other non-fentanyl opioids, as they appear in the marketplace.

Rapid screening of 2‐benzylbenzimidazole nitazene analogs in suspect counterfeit tablets using Raman, SERS, DART‐TD‐MS, and FT‐IR

“…2,3 In the last 3 years, there have been increased public health alerts from the forensic community and published reports on the emergence and abuse of several 2-benzylbenzimidazole compounds that are structurally different from opioids such as morphine and fentanyl but are thought to have similar or elevated potency relative to fentanyl. 4- 10 A review by Ujváry et al details the pharmacological properties of several 2-benzylbenzimidazole analgesics that were initially synthesized and patented in the 1950s but were never approved for use in the United States. 11 Several of these analgesic opioids, such as etonitazene, clonitazene, metonitazene, and isotonitazene, are listed as Schedule I controlled drugs under the United Nations Single Convention on Narcotic Drugs of 1961.…”

“…11,21 Several reports of overdose deaths in the United States have been linked to the emergence of these compounds, including isotonitazene, protonitazene, etonitazene, metonitazene, etodesnitazene, and N-piperidinyl etonitazene. [4][5][6][7][8][9][10][11]22,23 Tablets manufactured to look like Food and Drug Administration (FDA)-approved oxycodone HCl 30 mg tablets and those that are round, blue, half-scored, and debossed with "M" on the unscored side and "30" on the scored side (Supporting Information Figure S1) have been found to contain heroin, fentanyl, and fentanyl analogs using traditional laboratory analyses by FDA's Forensic Chemistry Center (FCC)…”

Identification of etazene (etodesnitazene) metabolites in human urine by LC‐HRMS

“…Etazene was found to be highly active at the μ‐opioid receptor (MOR) with a comparable efficacy as that of fentanyl and a potency exceeding that of morphine 4 . Etonitazene and N ‐pyrrolidino etonitazene are considered to be the most potent nitazene analogs so far 5 . Furthermore, etazene was also found to induce severe dose‐dependent toxicity in Danio rerio larvae, including increase in mortality, developmental malformations, and serious cardiotoxicity 6 …”

Notes from the Field: Nitazene-Related Deaths — Tennessee, 2019–2021

“…Nitazenes were created as a potential pain reliever medication nearly 60 years ago but have never been approved for use in the United States ( 2 ). Laboratory test results indicate that the potency of certain nitazene analogs (e.g., isotonitazene, protonitazene, and etonitazene) greatly exceeds that of fentanyl, whereas the potency of the analog metonitazene is similar to fentanyl ( 3 , 4 ). Naloxone has been effective in reversing nitazene-involved overdoses, but multiple doses might be needed ( 3 , 4 ).…”

“…Laboratory test results indicate that the potency of certain nitazene analogs (e.g., isotonitazene, protonitazene, and etonitazene) greatly exceeds that of fentanyl, whereas the potency of the analog metonitazene is similar to fentanyl ( 3 , 4 ). Naloxone has been effective in reversing nitazene-involved overdoses, but multiple doses might be needed ( 3 , 4 ). The prevalence of nitazene deaths in the United States is unknown and the frequency of nitazene involvement in overdose deaths in Tennessee has not yet been assessed.…”