A-series agent A-234: initial in vitro and in vivo characterization.

A-series agent A-234 belongs to a new generation of nerve agents. The poisoning of a former Russian spy Sergei Skripal and his daughter in Salisbury, England, in March 2018 led to the inclusion of A-234 and other A-series agents into the Chemical Weapons Convention. Even though five years have already passed, there is still very little information on its chemical properties, biological activities, and treatment options with established antidotes. In this article, we first assessed A-234 stability in neutral pH for subsequent experiments. Then, we determined its inhibitory potential towards human recombinant acetylcholinesterase (HssAChE; EC 3.1.1.7) and butyrylcholinesterase (HssBChE; EC 3.1.1.8), the ability of HI-6, obidoxime, pralidoxime, methoxime, and trimedoxime to reactivate inhibited cholinesterases (ChEs), its toxicity in rats and therapeutic effects of different antidotal approaches. Finally, we utilized molecular dynamics to explain our findings. The results of spontaneous A-234 hydrolysis showed a slow process with a reaction rate displaying a triphasic course during the first 72 h (the residual concentration 86.2%). A-234 was found to be a potent inhibitor of both human ChEs (HssAChE IC50 = 0.101 ± 0.003 µM and HssBChE IC50 = 0.036 ± 0.002 µM), whereas the five marketed oximes have negligible reactivation ability toward A-234-inhibited HssAChE and HssBChE. The acute toxicity of A-234 is comparable to that of VX and in the context of therapy, atropine and diazepam effectively mitigate A-234 lethality. Even though oxime administration may induce minor improvements, selected oximes (HI-6 and methoxime) do not reactivate ChEs in vivo. Molecular dynamics implies that all marketed oximes are weak nucleophiles, which may explain the failure to reactivate the A-234 phosphorus-serine oxygen bond characterized by low partial charge, in particular, HI-6 and trimedoxime oxime oxygen may not be able to effectively approach the A-234 phosphorus, while pralidoxime displayed low interaction energy. This study is the first to provide essential experimental preclinical data on the A-234 compound.

Efficacy of retigabine in ameliorating the brain insult following sarin exposure in the rat.

BACKGROUND: Sarin is an irreversible organophosphate cholinesterase inhibitor. Following toxic signs, an extensive long-term brain damage is often reported. Thus, we evaluated the efficacy of a novel anticonvulsant drug retigabine, a modulator of neuronal voltage gated K+ channels, as a neuroprotective agent following sarin exposure. METHODS: Rats were exposed to 1 LD50 or 1.2 LD50 sarin and treated at onset of convulsions with retigabine (5 mg/kg, i.p.) alone or in combination with 5 mg/kg atropine and 7.5 mg/kg TMB-4 (TA) respectively. Brain biochemical and immunohistopathological analyses were processed 24 h and 1 week following 1 LD50 sarin exposure and at 4 weeks following exposure to 1.2 LD50 sarin. EEG activity in freely moving rats was also monitored by telemetry during the first week following exposure to 1.2 LD50 and behavior in the Open Field was evaluated 3 weeks post exposure. RESULTS: Treatment with retigabine following 1 LD50 sarin exposure or in combination with TA following 1.2 LD50 exposure significantly reduced mortality rate compared to the non-treated groups. In both experiments, the retigabine treatment significantly reduced gliosis, astrocytosis and brain damage as measured by translocator protein (TSPO). Following sarin exposure the combined treatment (retigabine+ TA) significantly minimized epileptiform seizure activity. Finally, in the Open Field behavioral test the non-treated sarin group showed an increased mobility which was reversed by the combined treatment. CONCLUSIONS: The M current modulator retigabine has been shown to be an effective adjunct therapy following OP induced convulsion, minimizing epileptiform seizure activity and attenuating the ensuing brain damage.

Understanding the Interaction Modes and Reactivity of Trimedoxime toward MmAChE Inhibited by Nerve Agents: Theoretical and Experimental Aspects.

Organophosphorus (OP) compounds are used as both chemical weapons and pesticides. However, these agents are very dangerous and toxic to humans, animals, and the environment. Thus, investigations with reactivators have been deeply developed in order to design new antidotes with better efficiency, as well as a greater spectrum of action in the acetylcholinesterase (AChE) reactivation process. With that in mind, in this work, we investigated the behavior of trimedoxime toward the Mus musculus acetylcholinesterase (MmAChE) inhibited by a range of nerve agents, such as chemical weapons. From experimental assays, reactivation percentages were obtained for the reactivation of different AChE-OP complexes. On the other hand, theoretical calculations were performed to assess the differences in interaction modes and the reactivity of trimedoxime within the AChE active site. Comparing theoretical and experimental data, it is possible to notice that the oxime, in most cases, showed better reactivation percentages at higher concentrations, with the best result for the reactivation of the AChE-VX adduct. From this work, it was revealed that the mechanistic process contributes most to the oxime efficiency than the interaction in the site. In this way, this study is important to better understand the reactivation process through trimedoxime, contributing to the proposal of novel antidotes.

Oxime K203: a drug candidate for the treatment of tabun intoxication.

For over 60 years, researchers across the world have sought to deal with poisoning by nerve agents, the most toxic and lethal chemical weapons. To date, there is no efficient causal antidote with sufficient effect. Every trialed compound fails to fulfil one or more criteria (e.g. reactivation potency, broad reactivation profile). In this recent contribution, we focused our attention to one of the promising compounds, namely the bis-pyridinium reactivator K203. The oxime K203 is very often cited as the best reactivator against tabun poisoning. Herein, we provide all the available literature data in comprehensive and critical review to address whether K203 could be considered as a new drug candidate against organophosphorus poisoning with the stress on tabun. We describe its development from the historical point of view and review all available in vitro as well as in vivo data to date. K203 is easily accessible by a relatively simple two-step synthesis. It is well accommodated in the enzyme active gorge of acetylcholinesterase providing suitable interactions for reactivation, as shown by molecular docking simulations. According to a literature survey, in vitro data for tabun-inhibited AChE are extraordinary. However, in vivo efficiency remains unconvincing. The K203 toxicity profile did not show any perturbations compared to clinically used standards; on the other hand versatility of K203 does not exceed currently available oximes. In summary, K203 does not seem to address current issues associated with the organophosphorus poisoning, especially the broad profile against all nerve agents. However, its reviewed efficacy entitles K203 to be considered as a backup or tentative replacement for obidoxime and trimedoxime, currently only available anti-tabun drugs.

Repetitive antidotal treatment is crucial in eliminating eye pathology, respiratory toxicity and death following whole-body VX vapor exposure in freely moving rats.

Exposure to the chemical warfare nerve agent VX is extremely toxic, causing severe cholinergic symptoms. If not appropriately treated, death ultimately ensues. Based on our previously described whole-body vapor exposure system, we characterized in detail the clinical outcome, including respiratory dynamics, typical of whole-body exposure to lethal doses of VX vapor in freely moving rats. We further evaluated the efficacy of two different antidotal regimens, one comprising a single and the other repeated administration of antidotes, in countering the toxic effects of the exposure. We show that a 15 min exposure to air VX concentrations of 2.34-2.42 mg/m3 induced a late (15-30 min) onset of obvious cholinergic signs, which exacerbated over time, albeit without convulsions. Marked eye pathology was observed, characterized by pupil constriction to pinpoint, excessive lacrimation with red tears (chromodacryorrhea) and corneal damage. Respiratory distress was also evident, characterized by a three-fourfold increase in Penh values, an estimate of lung resistance, and by lung and diaphragm histological damage. A single administration of TAB (the oxime TMB-4, atropine and the anticholinergic and antiglutamatergic benactyzine) at the onset of clinical signs afforded only limited protection (66% survival), with clinical deterioration including weight loss, chromodacryorrhea, corneal damage, increased airway resistance and late death. In contrast, a combined therapy of TAB at the onset of clinical signs and repeated administration of atropine and toxogonin (ATOX) every 3-5 h, a maximum of five i.m. injections, led to 100% survival and a prompt recovery, accompanied by neither the above-described signs of eye pathology, nor by bronchoconstriction and respiratory distress. The necessity of recurrent treatments for successful elimination of VX vapor toxicity strongly supports continuous penetration of VX following termination of VX vapor exposure, most likely from a VX reservoir formed in the skin due to the exposure. This, combined with the above-described eye and respiratory pathology and absence of convulsions, are unique features of whole-body VX vapor exposure as compared to whole-body vapor exposure to other nerve agents, and should accordingly be considered when devising optimal countermeasures and medical protocols for treatment of VX vapor exposure.

Effect of six oximes on acutely anticholinesterase inhibitor-induced oxidative stress in rat plasma and brain.

Beside the key inhibition of acetylcholinesterase (AChE), involvement of oxidative stress in organophosphate (OP)-induced toxicity has been supported by experimental and human studies. On the other hand, according to our best knowledge, possible antioxidant properties of oximes, the only causal antidotes to OP-inhibited AChE, have been examined only by a few studies. Thus, we have determined the effect of four conventional (obidoxime, trimedoxime, pralidoxime, asoxime) and two promising experimental oximes (K027, K203) on dichlorvos (DDVP)-induced oxidative changes in vivo. Wistar rats (5/group) were treated with oxime (5% LD50 i.m) immediately after DDVP challenge (75% LD50 s.c). Oxidative stress biomarkers were determined in plasma and brain 60 min after the treatment: prooxidative-superoxide anion (O2·-) and total oxidative status (TOS); antioxidative-superoxide dismutase (SOD), total thiol (SH) groups, total antioxidant status (TAS) and paraoxonase (PON1); tissue oxidative stress burden-prooxidative-antioxidative balance (PAB) and oxidative stress index (OSI); oxidative tissue damage-malondialdehyde (MDA) and advanced oxidation protein products (AOPP). All oximes were able to attenuate DDVP-induced oxidative stress in rat plasma and brain. Changes of determined parameters in brain were not as prominent as it was seen in plasma. Based on OSI, better abilities of oxime K027, K203 and obidoxime to maintain DDVP-induced oxidative stress in rat brain were shown as compared to trimedoxime, pralidoxime and asoxime. Oximes can influence the complex in vivo redox processes that might contribute to their overall therapeutic efficacy. Further research is needed to understand the underlying molecular mechanisms involved in this phenomenon.

The Evaluation of the Reactivating and Neuroprotective Efficacy of Two Newly Prepared Bispyridinium Oximes (K305, K307) in Tabun-Poisoned Rats-A Comparison with Trimedoxime and the Oxime K203.

The ability of two newly developed oximes (K305, K307) to protect tabun-poisoned rats from tabun-induced inhibition of brain acetylcholinesterase, acute neurotoxic signs and symptoms and brain damage was compared with that of the oxime K203 and trimedoxime. The reactivating and neuroprotective effects of the oximes studied combined with atropine on rats poisoned with tabun at a sublethal dose were evaluated. The reactivating efficacy of a newly developed oxime K305 is lower compared to the reactivating efficacy of the oxime K203 and trimedoxime while the ability of the oxime K307 to reactivate tabun-inhibited acetylcholinesterase (AChE) in the brain roughly corresponds to the reactivating efficacy of the oxime K203 and it is slightly lower compared to trimedoxime. In addition, only one newly developed oxime (K307) combined with atropine was able to markedly decrease tabun-induced neurotoxicity although it did not eliminate all tabun-induced acute neurotoxic signs and symptoms. These results correspond to the histopathological evaluation of tabun-induced brain damage. Therefore, the newly developed oximes are not suitable for the replacement of commonly used oximes (especially trimedoxime) in the treatment of acute tabun poisonings.

A comparison of neuroprotective efficacy of two novel reactivators of acetylcholinesterase called K920 and K923 with the oxime K203 and trimedoxime in tabun-poisoned rats.

The ability of two newly developed bispyridinium oximes (K920, K923) to reduce tabun-induced acute neurotoxic signs and symptoms was compared with the oxime K203 and trimedoxime using a functional observational battery (FOB). The neuroprotective effects of the oximes studied combined with atropine on rats poisoned with tabun at a sublethal dose (130 µg/kg i.m.; 80% of LD50 value) were evaluated. Tabun-induced neurotoxicity was monitored by FOB at 2 h after tabun administration. The results indicate that all tested oximes combined with atropine enable tabun-poisoned rats to survive till the end of experiment while one non-treated tabun-poisoned rat died within 2 h. Both newly developed oximes (K920, K923) combined with atropine were able to markedly decrease tabun-induced neurotoxicity in the case of sublethal poisoning although they did not eliminate all tabun-induced acute neurotoxic signs and symptoms. Their ability to decrease tabun-induced acute neurotoxicity did not prevail the neuroprotective efficacy of trimedoxime and the oxime K203. Therefore, the newly developed oximes are not suitable for the replacement of currently available oximes (especially trimedoxime) in the treatment of acute tabun poisonings.

The Evaluation of the Potency of Newly Developed Oximes (K727, K733) and Trimedoxime to Counteract Acute Neurotoxic Effects of Tabun in Rats.

AIM: The ability of two newly developed oximes (K727, K733) to reduce tabun-induced acute neurotoxic signs and symptoms was evaluated and compared with currently available trimedoxime in rats. METHODS: The neuroprotective effects of the oximes studied combined with atropine on Wistar rats poisoned with tabun at a lethal dose (380 µg/kg i.m.; 90% of LD50 value) were evaluated. Tabun-induced neurotoxicity was monitored by the functional observational battery consisting of 38 measurements of sensory, motor and autonomic nervous functions at 2 hours following tabun challenge. RESULTS: All tested oximes combined with atropine enable tabun-poisoned rats to survive till the end of experiment. Both newly developed oximes (K727, K733) combined with atropine were able to decrease tabun-induced neurotoxicity in the case of lethal poisoning although they did not eliminate all tabun-induced acute neurotoxic signs and symptoms. CONCLUSION: The ability of both novel bispyridinium oximes to decrease tabun-induced acute neurotoxicity was slightly lower than that of trimedoxime. Therefore, the newly developed oximes are not suitable for the replacement of commonly used oximes such as trimedoxime in the treatment of acute tabun poisonings.

In vitro reactivation kinetics of paraoxon- and DFP-inhibited electric eel AChE using mono- and bis-pyridinium oximes.

Oxime-assisted reactivation of organophosphate (OP)-inhibited acetylcholinesterase (AChE) is a crucial step in the post-inhibitory treatment of OP intoxication. The limited efficacy of oxime reactivators for all OP nerve agents and pesticides led to the development of various novel oximes and their thorough kinetic investigations. Hence, in the present investigation, we have tested 10 structurally different pyridinium oxime-based reactivators for their in vitro potency to reactivate paraoxon- and DFP-inhibited electric eel AChE. From structure activity relationship point of view, various oximes such as mono-quaternary (2-PAM, K100, K024) and bis-quaternary symmetric (obidoxime, TMB-4) and asymmetric (K027, K048, K203, K618, K628) oximes bearing different connecting linkers (oxybismethylene, trimethylene, propane, butane, butene, and xylene) have been studied. The observed kinetic data demonstrate that not only the position of oxime group is decisive for the increased reactivation ability of oximes, but the role of connecting linker is also significant. Oximes with aliphatic linkers are superior reactivators than the oximes with unsaturated and aromatic linkers. The optimal chain length for plausible reactivation ability for paraoxon- and DFP-inhibited AChE is 3 or 4 carbon-carbon connecting linker between prydinium rings.

The evaluation of the reactivating and therapeutic efficacy of two novel oximes (K361 and K378) in comparison with the oxime K203 and trimedoxime in tabun-poisoned rats and mice.

The potency of two newly developed oximes (K361 and K378) to reactivate tabun-inhibited cholinesterase and to reduce acute toxicity of tabun was compared with the oxime K203 and trimedoxime using in vivo methods. The study determining percentage of reactivation of tabun-inhibited diaphragm cholinesterase in poisoned rats showed that the reactivating efficacy of the oxime K378 is slightly lower than the reactivating potency of the oxime K203 and trimedoxime while the ability of the oxime K361 to reactivate tabun-inhibited cholinesterase is markedly lower compared with the oxime K203 and trimedoxime. In the brain, the potency of both newly developed oximes to reactivate tabun-inhibited cholinesterase was negligible. The therapeutic efficacy of both newly developed oximes roughly corresponds to their weak reactivating efficacy. Their potency to reduce acute toxicity of tabun was significantly lower compared with the oxime K203 as well as trimedoxime. In conclusion, the reactivating and therapeutic potency of both newly developed oximes does not prevail the effectiveness of the oxime K203 and trimedoxime and, therefore, they are not suitable for their replacement of commonly used oximes for the treatment of acute tabun poisoning.

Pharmacokinetic study of two acetylcholinesterase reactivators, trimedoxime and newly synthesized oxime K027, in rat plasma.

K027 [1-(4-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium)-propane dibromide] is a promising new reactivator of organophosphate- or organophosphonate-inhibited acetylcholinesterase (AChE) with low acute toxicity and broad spectrum efficacy. The aim of the present study was to compare the pharmacokinetics of both compounds. Male Wistar rats (body weight = 320 ± 10 g) were administered a single intramuscular dose of K027 (22.07 mg kg(-1)) and an equimolar dose of trimedoxime. Blood was collected at various time intervals until 180 min. Plasma samples were analyzed by reversed-phase HPLC with ultraviolet (UV) detection. The recovery of both oximes from the plasma was approximately 90% and a linear relationship (R(2) > 0.998) was observed between the peak areas and concentrations of calibrated standards in the range 1-100 µg ml(-1). Near-identical plasma profiles were obtained for both compounds. No differences were found in the mean ± SD values of C(max) (18.6 ± 2.5 vs 20.0 ± 6.3 µg ml(-1), P = 0.72) and AUC(0-180min) (2290 ± 304 vs 2269 ± 197 min µg ml(-1), P = 0.84). However, the percentage coefficient of variation of the first-order rate constant of absorption (k(a)) was 3-fold higher (P < 0.01) providing evidence for more erratic absorption of intramuscular trimedoxime as compared with K027. In conclusion, oxime K027 might have superior pK properties that may be translated in its faster absorption and subsequent tissue distribution.

The evaluation of the reactivating and therapeutic efficacy of three novel bispyridinium oximes (K454, K456, K458) in comparison with the oxime K203 and trimedoxime in tabun-poisoned rats and mice.

The potency of three newly developed bispyridinium compounds (K454, K456, K458) to reactivate tabun-inhibited acetylcholinesterase and reduce tabun-induced lethal toxic effects was compared with the oxime K203 and trimedoxime using in vivo methods. The study determining percentage of reactivation of tabun-inhibited diaphragm and brain acetylcholinesterase in poisoned rats showed that the reactivating efficacy of all newly developed oximes is comparable with K203 but lower than the reactivating potency of trimedoxime in diaphragm. In the brain, their potency to reactivate tabun-inhibited acetylcholinesterase is lower compared with trimedoxime and the oxime K203. All three newly developed oximes were also found to be relatively effective in reducing lethal toxic effects in tabun-poisoned mice. Their therapeutic efficacy is consistent with the therapeutic potency of the oxime K203. On the other hand, their potency to reduce acute toxicity of tabun is significantly lower compared with trimedoxime. In conclusion, the reactivating and therapeutic potency of all three newly developed oximes does not prevail the effectiveness of the oxime K203 and trimedoxime and, therefore, they are not suitable for their replacement of commonly used oximes for the treatment of acute tabun poisoning.

[Studying kinetics of oxime-induced reactivation of malathion-inhibited cholinesterase].

The kinetics of oxime-induced reactivation of malathion-inhibited cholinesterase has been experimentally studied in vitro. It is shown that oximes do not restore the activity of inhibited butyrylcholinesterase. Acetylcholinesterase reactivation peak (5-mins long) was found to take place upon introduction of dipyroxime (32.5%), pralidoxime (18%), carboxyme (16%) at a concentration of 2.5 x 10(-4) mol/l or toxogonine (26%) at a concentration of 5 x 10(-4) mol/l. Toxogonine demonstrated the maximum affinity to phosphorylated enzyme, while dipyroxime is characterized by a high reactivity with respect to oxime. Significant reactivating ability of these preparations (kR -2300 mol(-1) min(-1) makes them promising solution for the treatment of malathion intoxication.

Molecular modeling of Mannich phenols as reactivators of human acetylcholinesterase inhibited by A-series nerve agents.

The A-series is the most recent generation of chemical warfare nerve agents (CWA) which act directly on the inhibition of the human acetylcholinesterase (HssAChE) enzyme. These compounds lack accurate experimental data on their physicochemical properties, and there is no evidence that traditional antidotes effectively reactivate HssAChE inhibited by them. In the search for potential antidotes, we employed virtual screening, molecular docking, and molecular dynamics (MD) simulations for the theoretical assessment of the performance of a library of Mannich phenols as potential reactivators of HssAChE inhibited by the Novichok agents A-230, A-232, and A-234, in comparison with the commercial oximes pralidoxime (2-PAM), asoxime (HI-6), trimedoxime (TMB-4), and obidoxime. Following the near-attack conformation (NAC) approach, our results suggest that the compounds assessed would face difficulties in triggering the proposed nucleophilic in-line displacement mechanism. Despite this, it was observed that certain Mannich phenols presented similar or superior results to those obtained by reference oximes against A-232 and A-234 model, suggesting that these compounds can adopt more favourable conformations. Additional binding energy calculations confirmed the stability of the model/ligands complexes and the reactivating potential observed in the molecular docking and MD studies. Our findings indicate that the Mannich phenols could be alternative antidotes and that their efficacy should be evaluated experimentally against the A-series CWA.

The benefit of combination of oximes for the neuroprotective efficacy of antidotal treatment of sarin-poisoned rats.

The potency of the oxime HI-6 and two combinations of oximes (HI-6 + trimedoxime, HI-6 + K203) to reduce sarin-induced acute neurotoxic signs and symptoms was evaluated in this study. Sarin-induced neurotoxicity and the neuroprotective effects of atropine alone or in combination with HI-6 alone and HI-6 combined with trimedoxime or K203 in rats poisoned with sarin at a sublethal dose (108 µg/kg i.m.; 90% of LD(50) value) were monitored by a functional observatory battery (FOB) 24 h following sarin administration. The results indicate that both mixtures of oximes combined with atropine were able to survive sarin-poisoned rats 24 h following sarin administration while two non-treated sarin-poisoned rats and one sarin-poisoned rat treated with atropine alone or with atropine in combination with the oxime HI-6 died within 24 h following sarin poisoning. All types of antidotal treatment were able to decrease sarin-induced neurotoxic signs and symptoms but not completely. While atropine alone and atropine in combination with the oxime HI-6 were able to eliminate some sarin-induced neurotoxic signs and symptoms, the neuroprotective efficacy of both combinations of oximes with atropine was slightly higher. Thus, both tested combinations of oximes in combination with atropine bring a small benefit for the neuroprotective efficacy of antidotal treatment of acute sarin poisonings.

The ability of oxime mixtures to increase the reactivating and therapeutic efficacy of antidotal treatment of cyclosarin poisoning in rats and mice.

The reactivating and therapeutic efficacy of two combinations ofoximes (HI-6 + trimedoxime and HI-6 + K203) was compared with the effectiveness of antidotal treatment involving single oxime (HI-6, trimedoxime, K203) using in vivo methods. In vivo determined percentage of reactivation of cyclosarin-inhibited blood and tissue acetylcholinesterase in poisoned rats showed that the reactivating efficacy of both combinations of oximes is slightly higher than the reactivating efficacy of the most effective individual oxime in blood, diaphragm as well as in brain. Moreover, both combinations of oximes were found to be slightly more efficacious in the reduction of acute lethal toxic effects in cyclosarin-poisoned mice than the antidotal treatment involving single oxime. Based on the obtained data, we can conclude that the antidotal treatment involving chosen combinations of oximes brings a beneficial effect for its ability to counteract the acute poisoning with cyclosarin.

Theoretical assessment of the performances of commercial oximes on the reactivation of acetylcholinesterase inhibited by the nerve agent A-242 (novichok).

The nerve agents of the A-series are relatively recent chemical weapons with no antidote available yet. Once inside the human body, those chemicals act similarly to the classic nerve agents, by binding to the catalytic residue Serine 203 (Ser203) of human acetylcholinesterase (HssAChE) and thus preventing the proper function of this enzyme. However, there is no experimental evidence yet if the current antidotes for intoxication by nerve agents are also capable of restoring AChE inhibited by the nerve agents of the A-series. In order to launch some light on this issue, we used computational techniques (molecular docking, molecular dynamics and MM-PBSA interaction energy calculations) to assess the performances of the four currently available commercial oximes (2-PAM, HI-6, obidoxime and trimedoxime) when in contact with HssAChE inhibited by the agent A-242. Based on the near-attack conformation (NAC) criterion, our results suggest that the commercial oximes would have limited efficacy to reactivate the enzyme since they are not able to properly approach the adduct Ser203-A-242. Among those oximes, trimedoxime seems to be the most promising, since it showed lower values of energy in the MM-PBSA calculations, a higher stability inside the catalytic anionic center (CAS) of HssAChE, and was able to adopt a position closer to the NAC that could enable the reactivation mechanism.

A comparison of the neuroprotective efficacy of individual oxime (HI-6) and combinations of oximes (HI-6+trimedoxime, HI-6+K203) in soman-poisoned rats.

The ability of two combinations of oximes (HI-6+trimedoxime, HI-6+K203) to reduce soman-induced acute neurotoxic signs and symptoms was compared with the neuroprotective efficacy of the oxime HI-6 alone, using a functional observational battery. Soman-induced neurotoxicity and the neuroprotective effects of HI-6 alone and HI-6 combined with trimedoxime or K203 in rats poisoned with soman at a sublethal dose (90 µg/kg intramuscularly, i.m.; 80% of LD50 value) were monitored by the functional observational battery at 24 hours following soman administration. The results indicate that both tested oxime mixtures combined with atropine were able to allow soman-poisoned rats to survive 24 hours following soman challenge, while 4 nontreated soman-poisoned rats and 1 soman-poisoned rat treated with oxime HI-6 alone combined with atropine died within 24 hours following soman poisoning. While the oxime HI-6 alone combined with atropine treatment was able to eliminate a few soman-induced neurotoxic signs and symptoms, both oxime mixtures showed higher neuroprotective efficacy in soman-poisoned rats. Especially, the combination of HI-6 with trimedoxime was able to eliminate most soman-induced neurotoxic signs and symptoms and markedly reduce acute neurotoxicity of soman in rats. Thus, both tested mixtures of oximes combined with atropine were able to increase the neuroprotective effectiveness of antidotal treatment of acute soman poisonings, compared to the individual oxime.

A comparison of the reactivating and therapeutic efficacy of chosen combinations of oximes with individual oximes against VX in rats and mice.

The ability of 2 combinations of oximes (HI-6 + trimedoxime and HI-6 + K203) to reactivate VX-inhibited acetylcholinesterase and reduce acute toxicity of VX was compared with the reactivating and therapeutic efficacy of antidotal treatment involving a single oxime (HI-6, trimedoxime, K203) in rats and mice. Our results showed that the reactivating efficacy of both combinations of oximes studied in rats is significantly higher than the reactivating efficacy of all individual oximes in diaphragm and roughly corresponds to the most effective individual oxime in blood and brain. Both combinations of oximes were found to be more effective in the reduction of acute lethal toxicity of VX in mice than the antidotal treatment involving the most efficacious individual oxime although the difference is not significant. Based on the obtained data, we can conclude that the antidotal treatment involving the chosen combinations of oximes brings benefit for the reactivation of VX-inhibited acetylcholinesterase in rats and for the antidotal treatment of VX-induced acute poisoning in mice.