PSTR-431
A novel M1 selective
muscarinic antagonist (AD-229) reverses acute effects of organophosphates on
respiratory rate and intracranial EEG
J. Thinschmidt1,
S.W. Harden1, J.D. Talton2, C.J. Frazier1
1Dept. of Cellular and Systems Pharmacology, College of
Pharmacy, Univ. of Florida, Gainesville, FL; 2Alchem Inc., Alachua
FL
Effective medical
countermeasures (MCMs) for organophosphate (OP) poisoning remain a critical
need due to the severe and often rapid neurotoxic effects of OP exposure.
Traditional therapies are limited by off-target interactions and poor CNS
bioavailability, largely due to inadequate penetration of the blood-brain
barrier (BBB). To address these limitations, there is ongoing interest in
identifying novel acetylcholinesterase (AChE) reactivators, muscarinic
antagonists, and other agents with broad-spectrum efficacy and enhanced CNS
activity. Our group has previously established a rat basolateral amygdala (BLA)
brain slice assay to characterize the efficacy of such compounds in
counteracting acute toxicity from the OP analog 4-nitrophenyl isopropyl
methylphosphonate (NIMP) (Thinschmidt et al. 2022). To complement this ex vivo
approach, we developed an anesthetized in vivo model that enables simultaneous
assessment of the BLA EEG and respiratory responses following intravenous OP
challenge (Thinschmidt et al 2024). In this model, 0.5 mg/kg NIMP (IV & SQ)
reliably induces seizure-like discharges in the BLA and progressive respiratory
depression. When administered intravenously, test compounds can be evaluated
for CNS efficacy based on their ability to mitigate these effects, thereby
providing insight into BBB permeability and pharmacodynamic action. Using this
platform, we previously demonstrated that intravenous administration of
atropine (5 mg/kg), a non-selective mAChR antagonist, effectively prevented and
reversed NIMP-induced seizure activity and respiratory depression. In contrast,
the M1-selective antagonist pirenzepine failed to provide protection or
reversal, consistent with its poor CNS penetration. Here, we evaluated AD-229,
a novel M1-selective muscarinic antagonist currently in preclinical
development, with an in vitro IC₅₀ of 16–32 nM. In both intravenous and
subcutaneous NIMP challenge models (0.5 mg/kg), AD-229 rapidly reversed BLA EEG
hyperactivity and respiratory depression, restoring both measures to
near-baseline levels. These results align with pharmacokinetic data showing a
half-life of 2.4 hours and robust CNS penetration, reflected in high
brain-to-plasma partitioning. Additional preclinical characterization of AD-229
reveals favorable properties, including an LD₅₀ > 50 mg/kg (single-dose,
mice) and aqueous formulation stability exceeding four months at room
temperature. Although GLP-compliant toxicity studies are pending, the current
data strongly support AD-229 as a promising lead compound with potential utility
as an alternative or adjunct to atropine in the treatment of OP poisoning.
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