Cholinesterase reactivation in vivo with a novel bis-oxime optimized by computer-aided design

Recently, several bis-pyridiniumaldoximes linked by a variable-length alkylene chain were rationally designed in our laboratories as cholinesterase reactivators. Extensive in vitro tests of these oximes with acetylcholinesterase inhibited by two different organophosphate agents, echothiophate and diisopropylfluorophosphate, revealed one compound with particularly good reactivation kinetics and affinity for phosphorylated acetylcholinesterase (AChE). This compound, designated "ortho-7", with a heptylene chain bridging two aldoximes ortho to a pyridinium ring nitrogen, was chosen for detailed comparison with the classic reactivator pyridine-2-aldoxime methochloride (2-PAM). In vitro, ortho-7 reactivated AChE selectively, without restoring activity of the related enzyme butyrylcholinesterase (BChE). For in vivo studies, rats were injected with ortho-7 or 2-PAM before or after organophosphate exposure, and the activities of AChE and BChE were determined at multiple intervals in blood and solid tissues. Ortho-7 behaved nearly as well in the animal as in vitro, reactivating AChE to the same extent as 2-PAM in all peripheral tissues studied (serum, red blood cell, and diaphragm), but at doses up to 100-fold smaller. Like other oxime reactivators, ortho-7 did not reactivate brain AChE after systemic administration. Nonetheless, this agent could be useful in combination therapy for organophosphate exposure, and it may provide a platform for development of additional, even more effective reactivators.