Outlining the A-Series of Organophosphorus Compounds: Cholinesterase Inhibition, Reactivation, Cytotoxicity, and Acute Toxicity in Mice.

Given the scarcity of experimental studies on A-series nerve agents (NAs), this paper provides ground-breaking insights and, for the first time, describes the inhibition and reactivation of human acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibited by these NAs using standard oximes. Furthermore, we present a detailed assessment of the toxicity profile of A-series NAs, based on both in vitro and in vivo studies. Our findings demonstrate that A-230, A-232, and A-234 are the most potent inhibitors of AChE and BChE among the A-series, with inhibitory potency comparable to the G-series NAs cyclosarin and soman. A-242 and A-262 inhibited both enzymes with potency similar to that of tabun and VX. Reactivation assays identified HI-6 oxime as the most effective reactivator in mitigating A-230-AChE conjugate-induced toxicity, achieving complete restoration of AChE activity within 4 hours. Obidoxime and TMB-4 showed moderate reactivation capacity over 24 hours, while 2-PAM was ineffective, indicating limited reactivation potential for counteracting A-230-inhibited AChE. These results confirm that both inhibition and reactivation are finely tuned processes, dependent on the structural characteristics of all reactants. In other words, while standard oximes show reasonable potency in reactivating AChE inhibited by agents like sarin and VX, they lack universal efficacy across different NA-AChE conjugates. The reactivation of BChE inhibited by A-agents was negligible when using standard oximes. Additionally, we modelled a near-attack conformation of HI-6 within AChE phosphylated by A-230, providing insights into the structural requirements necessary for more effective reactivation. Beyond enzyme studies, we also assessed hepatotoxicity and neurotoxicity in vitro, and evaluated the acute subcutaneous toxicity of A-series agents in mice. The toxicity of A-230, A-232, and A-234 was comparable to VX, while A-242 and A-262 were similar in toxicity to sarin. These findings significantly advance our understanding of the toxicological properties of phosphoramidates and underscore the urgent need to develop more effective medical countermeasures against A-agents exposure.