No description available.

Organophosphorus compounds (OPs) are used as pesticides, e.g. parathion, which is converted in the body to paraoxon, and chemical warfare nerve agents, such as sarin, soman, cyclosarin, VX, and tabun. Even small amounts of OP exposure can be fatal, depending on the toxicity of the compound. Great stocks of highly toxic chemical warfare nerve agents exit around the world and are considered a serious threat to national security and international stability. OPs exert their toxicity by covalent irreversible inhibition of acetylcholinesterase (AChE) that prevents the enzyme from hydrolyzing acetylcholine (ACh), a neurotransmitter in the central and peripheral nervous systems (CNS and PNS). Therefore, ACh accumulates in the cholinergic synapses throughout the body, which results in overstimulation of the ACh receptors. Removal of the phosphyl moiety from the OP-bound AChE active site has been a promising method to restore AChE’s catalytic activity. However, a secondary process called aging also occurs in the OP-AChE complex. Once aging occurs, currently available oximes are ineffective in removing the phosphyl moiety from the enzyme’s active site, and hence are ineffective as antidotes against the aged enzyme. Several families of alkylating and acylating agents including several classes of agents that combine alkylating moieties with known active site or peripheral cite (PAS) binding motifs were synthesized and evaluated. The general aim of the research was that successful alkylation or acylation of the phosphonate monoanion of aged AChE would produce neutral phosphyl complexes that would either spontaneously reactivate or would be reactivatable in the presence of oxime antidotes. Methoxylamine analogs of the oxime antidote 2-PAM were synthesized with the aim that methyl transfer to the aged AChE adduct would produce a neutral phosphyl AChE adduct simultaneously with 2-PAM in situ, and subsequent 2-PAM nucleophilic attack would reactivate the newly formed neutral phosphyl-AChE adduct. However, none of these 2-PAM analogs resurrected the activity of aged AChE. Another strategy for resurrecting the activity of aged AChE utilizes N-methylpyridiniums that are substituted at the 2-position with a beta-lactam moiety. For these compounds, opening of the electrophilic beta-lactam unmasks a nucleophilic amidine function which could putatively attack at phosphorus to expel the free enzyme. For this class of agents, only the active site directed compound that possessed the 5-CF₃ substituent showed possible resurrection of the activity of aged AChE, though activities in both the control and treated samples were low. Methyl transfers are common in Nature, and the natural transfer agent is S-adenosylmethionine, a sulfonium methyl donor. Consequently, the array of sulfonium compounds were evaluated on the expectation that they would bind to the AChE active site and transfer a methyl group to the phosphonate monoanion of the aged enzyme. Though high-affinity binding was noted for these compounds, none of these resurrected the activity of the aged AChE complex. Finally, several selected agents were evaluated on reactivating the initial OP-AChE complex before aging has occurred. It was observed that degraded samples of selected inhibitors are capable of reactivating initial complexes of sarin and soman inhibited AChE at low concentration that is an important character of efficient reactivators. However, the structure of reactivator is still unknown. Two major challenges still face researchers in the quest to design effective medicinal agents for counteracting poisoning by AChE-inhibiting nerve agents. The first is that there is no universal oxime antidote. Oximes that are effective against certain nerve agents are ineffective against others. The second is that, despite extensive efforts that span two generations, aged phosphyl-AChE adducts have never been reactivated. However, given the powerful tools of modern structural biology, medicinal chemistry and molecular biology, there is still hope that these considerable challenges can be met.

The aim of this study was to determine the ability of twenty novel substituted phenoxyalkyl pyridinium oximes to reactivate phosphylated acetylcholinesterase (AChE) in peripheral rat tissues, in vitro, inhibited by organophosphate anticholinesterase nerve agent surrogates. A sarin surrogate, phthalimidyl isopropyl methylphosphonate (PIMP), and a VX surrogate, 4-nitrophenyl ethyl methylphosphonate (NEMP), were used to inhibit AChE in skeletal muscle and serum samples. Reactivation of the widely used oxime 2-PAM was tested for comparison with the novel oximes. The novel oximes displayed a range of 23-102% reactivation of AChE in vitro across both tissue types. Most of the novel oximes tested in the present study demonstrated a higher percent reactivation of AChE, than 2-PAM. Therefore, these novel oximes have the potential to be effective antidotes used during the treatment of OP toxicity.

No description available.

Organophosphates (OPs), such as nerve agents and insecticides, potently inhibit acetylcholinesterase (AChE). Oximes, such as the currently FDA approved oxime 2-PAM, remove the OP from the inhibited enzyme. 2-PAM is effective against select OPs and cannot effectively pass the blood-brain barrier to attenuate OP induced CNS damage. Our laboratory has synthesized a series of substituted phenoxyalkyl pyridinium oximes (Patent number: 9,227,937) that have demonstrated increased survival rates compared to 2-PAM. This research investigated 1) in vitro oxime reactivation of rat, human, and guinea pig serum BChE after inhibition by nerve agent and insecticidal OPs; 2) in vitro determination of reactivation kinetic rate constants for OP inhibited human and rat serum BChE and electric eel AChE after inhibition by a sarin surrogate and paraoxon; 3) intranasal delivery of oximes to reactivate brain AChE in vivo after inhibition by a sarin surrogate. Novel oxime 15 demonstrated significant broad spectrum reactivation of OP-inhibited rat serum BChE while novel oxime 20 demonstrated significant broad spectrum reactivation of OP-inhibited human serum BChE. All tested oximes were poor reactivators of OP-inhibited guinea pig serum BChE. Kinetic analysis of reactivation for NIMP and paraoxon human and rat serum BChE and electric eel AChE demonstrated differences in the second order rate constants. Oxime 20 demonstrated reactivation efficiency for both NIMP and paraoxon inhibited rat and human serum BChE and electric eel AChE more effectively than 2-PAM. Intranasal delivery of either oxime 20 or 2-PAM showed attenuation of NIMP-inhibited brain AChE inhibition in select brain regions and select time points. Oxime 20 demonstrated a larger window of effectiveness but neither oxime attenuated brain AChE inhibition in the hindbrain for any time point or for any brain region at the ten minute time point. These data suggest that reactivation of OP-inhibited BChE may be contributing to the observed increases in survival seen with our oximes. Novel oxime 20 demonstrated reactivation efficacy towards both BChE and AChE inhibited enzyme and a rapid entry into the brain after intranasal delivery. Having an oxime that can be effective in a multitude of ways would be of great value to medical and military personnel.

The aim of this research was to determine whether a series of novel pyridinium oximes that were synthesized to incorporate phenoxyalkyl moieties to increase lipophilicity, and thereby increase their likelihood of crossing the blood-brain barrier, could effectively reactivate phosphorylated AChE in vitro. An experimental OP was synthesized as a surrogate for sarin to test the reactivation potential of the oximes, phthalimidyl isopropyl methylphosphonate (PIMP). The reactivation activities of the novel pyridinium oximes on PIMP exposed AChE and structure activity relationships were examined. Differences in reactivation potential in comparison to the widely used 2-PAM were also examined. All the novel oximes tested demonstrated some ability to reactivate inhibited AChE. Percent reactivations varied among the oximes (24%-78%), and the novel oximes were not as effective as 2-PAM or TMB-4, which reactivated 91 and 97%, respectively. The lipophilicity for all oximes was greater than 2-PAM or TMB-4 by 3- to 374- fold. A few of the novel oximes showed combined higher lipophilicity and reactivation potential approaching that of 2-PAM, and therefore suggest some potential efficacy as brain-penetrating oxime reactivators.

No description available.

Universiẗat, Diss., 2003--Dortmund.

Organophosphates (OPs), including nerve agents, target the cholinergic system via inhibition of acetylcholinesterase (AChE), with subsequent overstimulation resulting in neural damage and potential detrimental long-term effects. The efficacy of novel pyridinium oxime reactivators, created with moieties to increase blood-brain barrier penetration, was tested using highly relevant sarin and VX surrogates. Glial fibrillary acidic protein (GFAP; an indicator of neural damage) and monoamines (dopamine, serotonin, and their metabolites) were measured in select brain regions via immunohistochemistry and HPLC, respectively. Adult male rats were treated ip with high, sub-lethal doses of surrogates for sarin or VX, nitrophenyl isopropyl methylphosphonate (NIMP) or nitrophenyl ethyl methylphosphonate (NEMP), respectively. Surrogate treatment was followed after 1 hr by im administration of novel oxime. Seizure activity was monitored, and kainic acid (KA) served as a positive control. Administration of KA or surrogate (NIMP or NEMP) significantly increased GFAP expression compared to control animals. Two different formulations of one particular oxime (bromide vs. mesylate salt) attenuated seizures and reduced GFAP levels over NIMP or NEMP treatments alone to levels near those of controls in both the piriform cortex and dentate gyrus region of the hippocampus, while 2-PAM did not provide protection. Serotonergic activity was increased in several brain regions, including the piriform cortex, one hr after NIMP treatment. Markers of oxidative stress (isoprostanes) were also tested. Overall, these results indicate the potential therapeutic efficacy of these oximes and suggest this innovative chemistry may protect against neural damage induced by OP.

No description available.