The inhibition of acetylcholinesterase (AChE) by organophosphorus (OP) nerve agents and pesticides is, thankfully, reversible with the treatment of reactivators, such as 2-pralidoxime (2-PAM). However, if treatment is not administered quickly or if the OP is particularly toxic, these reactivators are rendered useless. After inhibition, a subsequent dealkylation event can occur at the phosphylated, catalytic serine residue of AChE. This aged state of the enzyme cannot be revived by reactivators, so the enzyme has been declared dead. There are currently no approved treatments for the aged form of AChE, and it was considered irreversible until 2018. Our team has demonstrated the only compounds that are capable of reviving, or resurrecting, the aged form of electric-eel AChE using quinone methide precursors (QMPs). Inspired by this initial discovery, a variety of QMPs have been tested <i>in vitro</i> against aged forms of recombinant human AChE that resemble the erythrocyte (dimer), brain (tetramer), and readthrough (monomer) isoforms. A modified Ellman’s assay, utilizing the artificial AChE substrate acetylthiocholine (ATC) and 5,5-dithio-bis-(2-nitrobenzoic acid) (DTNB), was used to detect the reappearance of native AChE during these studies. After 24 hours, up to 100% of the aged enzyme was resurrected for each enzyme variant with select QMPs and also for specific OP compounds. The same QMP framework is also an effective treatment for OP-inhibited AChE and can reactivate up to 70% of recombinant human (erythrocyte) AChE and 20% of the readthrough isoform. Although these QMPs are not as efficient with reactivation as 2-PAM, their lack of a permanent charge makes them more efficient at crossing the blood-brain barrier. QMPs, acting as both a reactivator and a resurrector, will help pave the way to new, more effective treatments for OP poisoning. In this presentation, we will present comparative <i>in vitro</i> assays with different variants of AChE as well as the resurrection efficiency with various QMPs in our structure-activity relationship library.

Resurrecting the dead: Recovery of organophosphorus poisoned acetylcholinesterase using quinone methide precursors

ACS Spring 2022

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