When a Prophecy comes True: Ethyleneoxynitazene as a ‘Prophetic’ Member of the Emerging Class of 2-Benzylbenzimidazole ‘Nitazene’ Synthetic Opioids

2-Benzylbenzimidazole opioids (‘nitazenes’) have become increasingly prevalent on the recreational drug market. We performed pharmacological characterization of various 'prophetic' nitazenes, allowing risk prioritization based on structure-activity relationships. Ethyleneoxynitazene, which we predicted to emerge and which was first found in January 2023, is presented as a case example. In vitro pharmacological characterization encompassed radioligand binding assays in rat brain tissue and a cell-based µ-opioid receptor activation (MOR-β-arrestin2) assay. Antinociception (hot plate assay), locomotor activity, and thermic effects were evaluated after subcutaneous administration in C57BL/6J mice. Binding assays revealed a Ki of 57.9 nM; only slightly higher than that of etonitazene (38.4 nM). However, ethyleneoxynitazene had a >100-fold lower potency in the MOR-β-arrestin2 assay (EC50, ethyleneoxynitazene=70.0 nM; EC50, etonitazene=0.588 nM). Its efficacy (relative to the reference hydromorphone) was also lower (Emax,ethyleneoxynitazene=187% vs. Emax,etonitazene=254%). The strongly reduced activity was reflected in vivo, with an ED50, antinociception of 11.1 mg/kg and 0.0223 mg/kg for ethyleneoxynitazene and etonitazene, respectively. Hypothermia and locomotor assays revealed the same pattern. The a priori availability of pharmacological data upon the first emergence of ethyleneoxynitazene allowed to rapidly communicate that (compared to other nitazenes) this is likely not the opioid of highest concern. Similarly, pharmacological data for other anticipated nitazenes are readily available.