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The misuse of novichok agents in assassination attempts has been reported in the international media since 2018. These relatively new class of neurotoxic agents is claimed to be more toxic than the agents of the G and V series and so far, there is no report yet in literature about potential antidotes against them. To shed some light into this issue, we report here the design and synthesis of NTMGMP, a surrogate of A-242 and also the first surrogate of a novichok agent useful for experimental evaluation of antidotes. Furthermore, the efficiency of the current commercial oximes to reactivate NTMGMP-inhibited acetylcholinesterase (AChE) was evaluated. The Ellman test was used to confirm the complete inhibition of AChE, and to compare the subsequent rates of reactivation in vitro as well as to evaluate aging. In parallel, molecular docking, molecular dynamics and MM-PBSA studies were performed on a computational model of the human AChE (HssAChE)/NTMGMP complex to assess the reactivation performances of the commercial oximes in silico. Experimental and theoretical studies matched the exact hierarchy of efficiency and pointed to trimedoxime as the most promising commercial oxime for reactivation of AChE inhibited by A-242. [ABSTRACT FROM AUTHOR]

Organophosphorus poisoning caused by some pesticides and nerve agents is a life-threating condition that must be swiftly addressed to avoid casualties. Despite the availability of medical countermeasures, the clinically available compounds lack a broad spectrum, are not effective towards all organophosphorus toxins, and have poor pharmacokinetics properties to allow them crossing the blood-brain barrier, hampering cholinesterase reactivation at the central nervous system. In this work, we designed and synthesised novel isatin derivatives, linked to a pyridinium 4-oxime moiety by an alkyl chain with improved calculated properties, and tested their reactivation potency against paraoxon- and NEMP-inhibited acetylcholinesterase in comparison to the standard antidote pralidoxime. Our results showed that these compounds displayed comparable in vitro reactivation also pointed by the in silico studies, suggesting that they are promising compounds to tackle organophosphorus poisoning. [ABSTRACT FROM AUTHOR]

Organophosphates, useful agents as pesticides, also represent a serious danger due to their high acute toxicity. There is indication that oximes, when administered before organophosphate exposure, can protect from these toxic effects. We have tested at equitoxic dosage (25% of LD01) the prophylactic efficacy of five experimental (K‐48, K‐53, K‐74, K‐75, K‐203) and two established oximes (pralidoxime and obidoxime) to protect from mortality induced by the organophosphate paraoxon. Mortalities were quantified by Cox analysis and compared with those observed after pretreatment with a strong acetylcholinesterase inhibitor (10‐methylacridine) and after the FDA‐approved pretreatment compound pyridostigmine. All nine tested substances statistically significantly reduced paraoxon‐induced mortality. Best protection was conferred by the experimental oxime K‐48, reducing the relative risk of death (RR) to 0.10, which was statistically significantly superior to pyridostigmine (RR = 0.31). The other oximes reduced the RR to 0.13 (obidoxime), 0.20 (K‐203), 0.21 (K‐74), 0.24 (K‐75) and 0.26 (pralidoxime), which were significantly more efficacious than 10‐methylacridine (RR = 0.65). These data support the hypothesis that protective efficacy is not primarily due to cholinesterase inhibition and indicate that the tested experimental oximes may be considered promising alternatives to the established pretreatment compound pyridostigmine. The prophylactic efficacy of five experimental (K‐48, K‐53, K‐74, K‐75, K‐203) and two established oximes (pralidoxime and obidoxime) to protect from mortality induced by the organophosphate paraoxon was evaluated. Best protection was conferred by the experimental oxime K‐48, reducing the relative risk of death to 0.10, which was significantly superior to the FDA‐approved substance pyridostigmine. [ABSTRACT FROM AUTHOR]

In this study, it has been investigated the oxidative status and morphological alterations in the brain of Wistar rats induced by repeated application of low doses of selected acetylcholinesterase reactivators - asoxime, obidoxime, K027, K048, K074, and K075. Each oxime (0.1 of LD50/kg im) was given 2 times per week for 4 weeks. The animals' whole-body, organ weight, oxidative status, as well as microscopic examination of the brain, were done on day 35 of the study. Markers of oxidative stress, lipid peroxidation (malondialdehyde (MDA) and advanced oxidation protein products (AOPP)), as well as the activity of antioxidant enzymes (catalase (CAT) and superoxide dismutase (SOD)), were measured in the brain homogenates (1,2). Brain alterations were quantified by semiquantitative grading scales - brain damage score (BDS) (3). Oxidative stress parameters MDA and AOPP were significantly elevated in the K075-treated group compared to the control group (p < 0.001). SOD activity was significantly higher in the K075-treated group (p < 0.001), without differences in CAT activity. In the same groups of rats, brain injuries were significantly more severe than those observed in animals receiving only asoxime or K027 (p < 0.001). Our results can help to predict likely adverse systemic toxic effects, and target organ systems, which are crucial for establishing risk categories, as well as in dose selection of K-oximes as drug candidates. (MFVMA01/23-25). [ABSTRACT FROM AUTHOR]

Benchmark dose (BMD) approach, as an advanced statistical methodology for dose-effect analysis in toxicological research (1) was used to quantitatively characterize in vivo efficacy of two experimental bispyridinium oximes K203 and K027, following promising findings on their low acute toxicity and potential to reactivate acetylcholinesterase (AChE) inhibited by organophosphorus (OP) pesticides and nerve agents in vitro and in vivo (2). Immediately after DDVP challenge (75% LD50, s.c.), male Wistar rats were treated with oxime (0/1.25/2.5/5/25/50% LD50, i.m.). Erythrocyte and diaphragm AChE activity was determined by Ellman's method 60 min after the treatment. Benchmark analysis was done in PROAST software ver. 65.5 (RIVM, The Netherlands). Derived BMDer were K203 = 194 (153, 243) and K027 = 100 (81, 125) μmol/kg bw, BMDdiaph were K203 = 117 (56, 209) and K027 = 21 (10, 37) μmol/kg bw, indicating that oxime K027 induces the same effect size with 2 and 5.5-times lower dose compared to oxime K203 in erythrocites and diaphragm, respectively. Quantification of equieffective doses of oxime reactivators would enable more reliable definition of their therapeutic widths, which further contributes to better determination of therapeutic dosage regimens and, finally, increases the relevance of results obtained in animal models for the human population. [ABSTRACT FROM AUTHOR]

Copyright of Archives of Industrial Hygiene & Toxicology / Arhiv za Higijenu Rada I Toksikologiju is the property of Sciendo and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Pre-treatment with reversible acetylcholinesterase (AChE) inhibitors before organophosphorous compound (OPC) exposure can reduce OPC-induced mortality. However, pyridostigmine, the only substance employed for such prophylaxis, is merely efficacious against a limited number of OPCs. In search of more efficacious and broad-range alternatives, we have compared in vivo the ability of five reversible AChE inhibitors (pyridostigmine, physostigmine, ranitidine, tacrine and K-27) to reduce mortality induced by the OPC azinphos-methyl. Protection was quantified using Cox analysis by determining the relative risk (RR) of death in rats that were administered these AChE inhibitors in equitoxic dosage (25% of LD01) 30 min before azinphos-methyl exposure. Azinphos-methyl-induced mortality was significantly reduced by all five tested compounds as compared with the reference group that was only exposed to azinphos-methyl without prior pre-treatment (RR = 1). The most efficacious prophylactic agents were K-27 (RR = 0.15) and physostigmine (RR = 0.21), being significantly more efficacious than ranitidine (RR = 0.62) and pyridostigmine (RR = 0.37). Pre-treatment with tacrine (RR = 0.29) was significantly more efficacious than pre-treatment with ranitidine, but the difference between tacrine and pyridostigmine was not significant. Our results indicate that prophylactic administration of the oxime K-27 may be a promising alternative in cases of imminent OPC exposure. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

ABSTRACT Poisoning with organophosphorus compounds (OPCs) poses a serious threat worldwide. OPC-induced mortality can be significantly reduced by prophylactic administration of reversible acetylcholinesterase (AChE) inhibitors. The only American Food and Drug Administration (FDA)-approved substance for such pre-treatment (to soman exposure) is presently pyridostigmine, although its efficacy is controversial. In search for more efficacious and broad-spectrum alternatives, we have assessed in vivo the mortality-reducing efficacy of a group of five compounds with known AChE inhibitory activity (pyridostigmine, physostigmine, ranitidine, tacrine and K-27), when given in equitoxic dosage (25% of LD01) 30 min before exposure to the OPC terbufos sulfone. Protection was quantified in rats by determining the relative risk of death (RR) using Cox analysis, with RR = 1 for animals given only terbufos sulfone, but no pre-treatment. All tested AChE inhibitors reduced terbufos sulfone-induced mortality significantly (p ≤ 0.05) as compared with the non-treatment group (RR = 1: terbufos sulfone only). Best in vivo protection from terbufos sulfone-induced mortality was achieved, when K-27 was given before terbufos sulfone exposure (RR = 0.06), which was significantly ( P ≤ 0.05) superior to the pre-treatment with all other tested compounds, for example tacrine (RR = 0.21), pyridostigmine (RR = 0.28), physostigmine (RR = 0.29) and ranitidine (RR = 0.33). The differences in efficacy between tacrine, pyridostigmine, physostigmine and ranitidine were not statistically significant. Prophylactic administration of an oxime (such as K-27) in case of imminent OPC exposure may be a viable option. Copyright © 2013 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

K048 is a member of K-oximes, a new oxime class that has recently been confirmed effective against poisoning by the nerve agent tabun and several pesticides. The toxicity profile of the K048 oxime has not been fully characterized and its optimal therapeutic dose has not yet been established. Earlier studies report excellent results with K048 in reactivating tabun-phosphorylated AChE and in the therapy of tabun-poisoned mice. It possesses a low acute toxicity and exerts an acceptable toxicity profile on isolated human peripheral blood lymphocytes in vitro. Intraperitoneal administration of K048 in rats resulted in an LD50 of 238.3 mg/kg. In this in vivo study, we investigated cholinesterase ( ChE) activity and oxidative stress marker levels (lipid peroxidation and superoxide dismutase activity) in the plasma of exposed rats after administering the compound at 25% of its LD50. Lymphocyte viability was evaluated using an acridine orange/ethidium bromide in situ fluorescent assay. The levels of primary DNA damage in rat white blood cells were measured using the alkaline comet assay. The compound applied at 25% of its LD50 did not significantly affect ChE activity and lipid peroxidation and did not cause significant changes in the SOD activity in plasma. The cytotoxicity profile of K048 in the tested dose was also acceptable, and it did not possess significant DNA-damaging potential. The obtained results are promising for further evaluations of the K048 oxime, which should include tests on a broader concentration range and longer incubation times. [ABSTRACT FROM AUTHOR]

Summary The neuroprotective effects of a novel oxime KR-22934, the oxime K203 and obidoxime in combination with atropine in rats poisoned with tabun at a sublethal dose (200 μg/kg i.m.; 80% LD 50 ) were studied. The tabun-induced neurotoxicity was monitored at 24 h following tabun challenge using a functional observational battery and an automatic measurement of motor activity. The results indicate that all tabun-poisoned rats treated with oximes in combination with atropine were able to survive within 24 h following tabun poisoning. One tabun-poisoned rat without antidotal treatment died within 24 h. The oximes KR-22934 and K203 combined with atropine showed a similar potency to decrease tabun-induced neurotoxicity at 24 h after tabun administration while the neuroprotective efficacy of obidoxime was slightly higher. However, no oxime was able to eliminate tabun-induced neurotoxicity completely. When atropine was administered alone, negligible neuroprotective efficacy was observed. Based on the results, a novel oxime KR-22934 did not bring any improvement of the neuroprotective efficacy of antidotal treatment of acute tabun poisonings. [ABSTRACT FROM AUTHOR]

The chromatographic behavior was studied of a series of potential acetylcholinesterase reactivators, pyridinium oximes, bearing linear aliphatic chains of the length of the aliphatic bridge from 1 to 12 carbon atoms, on a pentafluorophenyl-modified stationary phase. The retention mechanisms and the dependence of the capacity factor on mobile phase composition, aliphatic chain bridge length and calculated log P were evaluated and discussed in detail. The separation of the studied oximes was found to be driven by hydrophobic interactions when a lower content of organic modifier was used in mobile phase; however, the ion-exchange mechanism was the leading one when a large portion of organic modifier was used. In addition, the lipophilicity was found to be a driving mechanism of the separation of oximes bearing a connecting chain of the length of 6–12 carbon atoms, whereas the retention of oximes with shorter connecting chains was significantly influenced by other separation mechanisms such as aromatic or π–π interaction. These results can be useful for the development of new, efficient acetylcholinesterase reactivators. [ABSTRACT FROM AUTHOR]

Acetylcholinesterase reactivators (oximes) are highly hydrophilic compounds. This factor is considered to limit their distribution through the human body, especially into the central nervous system. However, some results contradict this hypothesis and confirm low concentrations of oximes inside the CNS. This study aims to verify oxime penetration through the blood-brain barrier, and into different brain regions. It has been confirmed that oximes can penetrate the BBB after therapeutic dose administration (0.73%, 0.69%, and 0.52% corresponding to obidoxime, HI-6, and new oxime K027). Although the whole brain level of K027 seems to be lower compared with other oximes we have to keep in mind that these numbers represent only a percentage of the concentration found in plasma. So, for more adequate comparison it is essential to have a look at the actual concentration of oximes in the wet brain tissue: 106.6 ± 19.87 ng/g (K027), 49.88 ± 10.21 ng/g (obidoxime) and 86.51 ± 42.37 ng/g (HI-6), where concentration of K027 was one order of magnitude higher than obidoxime and HI-6. In spite of the fact that the distribution of oximes into the CNS is minimal and concentrations differ from section to section, reasonable reactivation should still occur. [ABSTRACT FROM AUTHOR]

ABSTRACT Reversible acetylcholinesterase (AChE) inhibitors can protect against the lethal effects of irreversible organophosphorus AChE inhibitors (OPCs), when administered before OPC exposure. We have assessed in vivo the mortality-reducing efficacy of a group of known AChE inhibitors, when given in equitoxic dosage before exposure to the OPC paraoxon. Protection was quantified in rats by determining the relative risk (RR) of death. Best in vivo protection from paraoxon-induced mortality was observed after prophylactic administration of physostigmine (RR = 0.30) or the oxime K-27 (RR = 0.34); both treatments were significantly superior to the pre-treatment with all other tested compounds, including the established substance pyridostigmine. Tacrine (RR = 0.67), ranitidine (RR = 0.72), pyridostigmine (RR = 0.76), tiapride (RR = 0.80) and 7-MEOTA (RR = 0.86) also significantly reduced the relative risk of paraoxon-induced death, but to a lesser degree. Methylene blue, amiloride and metoclopramide had an unfavorable effect (RR ≥ 1), significantly increasing mortality. When CNS penetration by prophylactic is undesirable K-27 is a promising alternative to pyridostigmine. Copyright © 2012 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Prophylactic administration of reversible acetylcholinesterase (AChE) inhibitors before exposure to organophosphorus compounds (OPCs) can reduce OPC-induced mortality. Pyridostigmine is the only FDA-approved substance for such use. The AChE-inhibitory activity of known AChE inhibitors was quantified in vitro and their in vivo mortality-reducing efficacy was compared, when given prophylactically before the exposure to the OPC diisopropylfluorophosphate (DFP). The IC50 was measured in vitro for the known AChE inhibitors pyridostigmine, physostigmine, ranitidine, tiapride, tacrine, 7-methoxytacrine, amiloride, metoclopramide, methylene blue and the experimental oxime K-27. Their in vivo efficacy, when given as pretreatment, to protect rats from DFP-induced mortality was quantified by determining the relative risk of death (RR) by Cox analysis, with RR = 1 for animals given only DFP, but no pretreatment. Physostigmine was the strongest in vitro AChE-inhibitor (IC50 = 0.012 µ m), followed by 7-methoxytacrine, tacrine, pyridostigmine and methylene blue. Ranitidine (IC50 = 2.5 µ m), metoclopramide and amiloride were in the mid-range. Tiapride (IC50 = 256 µ m) and K-27 (IC50 = 414 µ m) only weakly inhibited RBC AChE activity. Best in vivo protection from DFP-induced mortality was achieved when physostigmine (RR = 0.02) or tacrine (RR = 0.05) was given before DFP exposure, which was significantly superior to the pretreatment with all other tested compounds, except K-27 (RR = 0.18). The mortality-reducing effect of pyridostigmine, ranitidine and 7-methoxytacrine was inferior, but still significant. Tiapride, methylene blue, metoclopramide and amiloride did not significantly improve DFP-induced mortality. K-27 may be a more efficacious alternative to pyridostigmine, when passage into the brain precludes administration of physostigmine or tacrine. Copyright © 2010 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Organophosphorus compounds pose a potential threat to both military and civilian populations. Since post-exposure therapy has its limitations, our research was focused on the possibility of improving pretreatment in order to limit the toxic effects of tabun. We determined the protective index of various combinations of atropine, oximes (K074, K048, and TMB-4), and pyridostigmine given to mice before tabun intoxication. Although the tested oximes showed very good therapeutic efficacy in tabun-poisoned mice, the given pretreatments improved therapy against tabun poisoning. These regimens ensured survival of all animals up to 25.2 LD50 of tabun. Our results indicate that even pretreatment with atropine alone is sufficiently effective in enhancing the survival of mice poisoned by multiple doses of tabun, if oxime therapy follows. K048 is our oxime of choice for future research, as it shows better protective and reactivating potency. [ABSTRACT FROM AUTHOR]

The pyridinium oxime therapy for treatment of organophosphate poisoning is a well established, but not sufficient method. Recent trends also focus on prophylaxis as a way of preventing even the entrance of organophosphates into the nervous system. One of the possible prophylactic methods is increasing the concentration of butyrylcholinesterase in the blood with the simultaneous administration of butyrylcholinesterase reactivators, when the enzyme is continuously reactivated by oxime. This article summarizes and sets forth the structural differences between butyrylcholinesterase and acetylcholinesterase, essential for the future design of butyrylcholinesterase reactivators. Butyrylcholinesterase lacks the reactivator aromatic binding pocket found in acetylcholinesterase, which is itself a part of the acetylcholinesterase peripheral anionic site. This difference finally renders the current acetylcholinesterase reactivators, when used in butyrylcholinesterase, non-functional. [ABSTRACT FROM AUTHOR]

In this in vitro study, using the HPLC method, we determined the ability of acetylcholinesterase (AChE) reactivators, used clinically, to penetrate the blood-brain barrier (BBB). We evaluated pralidoxime, HI-6, obidoxime, trimedoxime and methoxime - reactivators varying in the position of the oxime group on the pyridinium ring and linker connecting the pyridinium rings. Our results indicated that pralidoxime, a monoquaternary AChE reactivator, was the oxime with the most penetration. Molecular weight seems to be the most important factor for passive transport through the BBB. From the structural perspective, the connecting linker also plays a key role in the ability of the reactivators to penetrate the CNS. In this case, the simple and short linker is favorable for permeation of these compounds. The location of the oxime group on the pyridine ring may also influence passive transport into the brain; the best position of the oxime group seems to be position four. [ABSTRACT FROM AUTHOR]

The broad-spectrum reactivator is a valuable oxime able to reactivate acetylcholinesterase (AChE) inhibited by nerve agents and pesticides. At present there are many AChE reactivators (oximes) which are suitable candidates as broad-spectrum reactivators and among them is the oxime HI-6, highly enough thought of to have been recommended by many armies for use as a universal antidote. In this study, we wanted to establish whether the designation "broad-spectrum" is a an accurate description or if there are some lacks in reactivation of nerve agents or pesticides. For this purpose, the general in vitro test for the evaluation ofAChE reactivators was used. Tabun, sarin, cyclosarin, soman, VX agent, Russian VX were used as nerve agents for testing, and chiorpyrifos, paraoxon, methyl-chiorpyrifos and dichlorvos (DDVP) were used as typical examples of organophoshorus pesticides. The results obtained showed that oxime HI-6 did .not reactivate tabun- and DDVP-inhibited AChE, and, in the case of the other pesticides, only a high dose of oxime HI-6 was able to reactivate pesticide-inhibited AChE. [ABSTRACT FROM AUTHOR]

Copyright of Archives of Industrial Hygiene & Toxicology / Arhiv za Higijenu Rada I Toksikologiju is the property of Sciendo and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The article presents an in vitro study of three potential acetylcholinesterase (AChE) reactivators taken from a monoquaternary reactivator pralidoxime. The compounds used in the study include pyridinium-2-aldoxime-4- carbamoyl-N-methyl iodide, pyridinium-2-aldoxime-4-ethoxycarbonyl-N-methyl iodide, and pyridinium-2-aldoxime-5-ethoxycarbonyl-N-methyl iodide. Results showed that none of the tested oximes was able to reactivate cyclosarin-inhibited AChE.

In this work in vitro evaluation of the reactivation potency of the newly synthesized reactivators for acetylcholinesterase (AChE; EC 3.1.1.7) is described. Using this method, reactivation potency of 21 potential reactivators of AChE inhibited by the nerve agent sarin has been evaluated. We have confirmed the fact that currently the most promising AChE reactivator, HI-6, is the most effective reactivator of sarin-inhibited AChE. There are only three AChE reactivators—HI-6, TO033 and TO047—able to satisfactorily reactivate sarin-inhibited AChE at the concentration 10 -5 M, which is nontoxic for human use. On the other hand, there are 14 AChE reactivators, that are able to reactivate sarin-inhibited AChE at the concentration 10 -3 M. However, this concentration of reactivator is probably toxic for human use. [ABSTRACT FROM AUTHOR]

Abstract: It was shown that intoxications with GF-agent are rather resistant to convential oxime therapy; therefore, the development of new oximes in an effort to improve this unsatisfactory situation continues. Upon screening in vitro reactivation test for oximes, that were either newly synthesized at our department, or those that have never been tested for reactivation of GF-inhibited acetylcholinesterase (AChE), three oximes {(1,4-bis(4-hydroxyiminomethylpyridinium)butane dibromide) (K033); (1-(2-hydroxyiminomethylpyridinium)-3-(3-carbamoylpyridinium)-2-oxa-propane dichloride) (HS-6); and (1-(2-hydroxyiminomethylpyridinium)-4-(4-carbamoylpyridinium)-but-2-ene dibromide) (BI-6)} with the highest reactivation potency were chosen for in vivo testing in our study. 1,3-Bis(4-hydroxyiminomethylpyridinium)-2-oxa-propane dibromide) (obidoxime); (1-(2-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium)-2-oxa-propane dichloride) (HI-6); and (1,1-bis(4-hydroxyiminomethylpyridinium)-methane dibromide) (methoxime) were chosen for comparison as a standard antidotal treatment. All the oximes were applied at the same proportion of their LD50 value (5%), and because of the different acute toxicity of the oximes, the molar concentrations of their solutions for intramuscular (i.m.) administration were considerably different. The highest therapeutic ratio was achieved for therapeutic regimen consisting of HI-6 and atropine. The significantly (P < 0.05) lowest effectivity in treatment of supralethal GF-agent poisoning in comparison with all the other therapeutic regimens, was surprisingly observed for methoxime. HS-6, K033 and BI-6 as well as obidoxime were comparably effective antidotes against GF-agent intoxication and their therapeutic ratios were similar. [Copyright &y& Elsevier]

A comparison of one mono- and seven bisquaternary acetylcholinesterase (AChE) reactivators of acetylcholinesterase inhibited by VX agent was performed. As a source of the acetylcholinesterase, a rat brain homogenate was taken. There were significant differences in reactivation potency of all tested oximes. The oxime TO205 seems to be the most efficacious followed by TO046, HI-6, HS-6, K027, obidoxime, MMC and 2-PAM. In addition, the results of this study showed that the reactivation potency of the tested reactivators depends on many factors - such as the number of pyridinium rings, the number of oxime groups and their position, as well as the length and the shape of linkage bridge between two pyridinium rings. [ABSTRACT FROM AUTHOR]

Aims: Organophosphates (OPCs), useful agents as pesticides, also represent a serious health hazard. Standard therapy with atropine and established oxime-type enzyme reactivators is unsatisfactory. Experimental data indicate that superior therapeutic results can be obtained when reversible cholinesterase inhibitors are administered before OPC exposure. Comparing the protective efficacy of five such cholinesterase inhibitors (physostigmine, pyridostigmine, ranitidine, tacrine, or K-27), we observed best protection for the experimental oxime K-27. The present study was undertaken in order to determine if additional administration of K-27 immediately after OPC (paraoxon) exposure can improve the outcome. Methods: Therapeutic efficacy was assessed in rats by determining the relative risk of death (RR) by Cox survival analysis over a period of 48 h. Animals that received only pretreatment and paraoxon were compared with those that had received pretreatment and paraoxon followed by K-27 immediately after paraoxon exposure. Results: Best protection from paraoxon-induced mortality was observed after pretreatment with physostigmine (RR = 0.30) and K-27 (RR = 0.34). Both substances were significantly more efficacious than tacrine (RR = 0.67), ranitidine (RR = 0.72), and pyridostigmine (RR = 0.76), which were less efficacious but still significantly reduced the RR compared to the no-treatment group (paraoxon only). Additional administration of K-27 immediately after paraoxon exposure (posttreatment) did not further reduce mortality. Statistical analysis between pretreatment before paraoxon exposure alone and pretreatment plus K-27 posttreatment did not show any significant difference for any of the pretreatment regimens. Conclusions: Best outcome is achieved if physostigmine or K-27 are administered prophylactically before exposure to sublethal paraoxon dosages. Therapeutic outcome is not further improved by additional oxime therapy immediately thereafter. [ABSTRACT FROM AUTHOR]

Certain AChE reactivators, asoxime, obidoxime, K027, K048, and K075, when taken in overdoses and sometimes even when introduced within therapeutic ranges, may injure the different organs. As a continuation of previously published data, in this study, Wistar rats have sacrificed 24 hrs and 7 days after single im application of 0.1LD50, 0.5LD50 and 1.0LD50 of each reactivator, and examinated tissue samples were obtained for pathohistological and semiquantitative analysis. A severity of tissue alteration, expressed as different tissue damage scores were evaluated. Morphological structure of examinated tissues treated with of 0.1LD50 of all reactivators was comparable with the control group of rats. Moderate injuries were seen in visceral tissues treated with 0.5LD50 of asoxime, obidoxime and K027. Acute damages were enlarged after treatment with 0.5LD50 and 1.0LD50 of all reactivators during the next 7 days. The most prominent changes were seen in rats treated with 1.0LD50 of K048 and K075 (P < 0.001 vs. control and asoxime-treated group). All reactivators given by a single, high, unitary dose regimen, have an adverse effect not only on the main visceral tissue, but on the whole rat as well, but the exact mechanism of cellular injury remains to be confirmed in further investigation. [ABSTRACT FROM AUTHOR]

Two methods for the synthesis of a new unsymmetric bispyridinium oxime-1-(4-hydroxyiminomethylpyridinium)-3-(carbamoylpyridinium)propane dibromide are described. In vitro efficacy of this new oxime to reactivate sarin-inhibited acetylcholinesterase has been evaluated. [Copyright &y& Elsevier]

Newly developed oximes, when taken in overdoses and sometimes even when introduced within therapeutic ranges, may injure the different organs. In this work, we focused our attention on an investigation of morphological lesions produced by increasing doses of asoxime, obidoxime, K027, K048, and K075 were selected as experimental reactivators. The whole experiment was conducted on Wistar rats. All rats were sacrificed 24 hrs and 7 days after single im application of 0.1 LD50, 0.5 LD50 and 1.0 LD50 of each reactivator. Tissue alterations were carefully quantified by semiquantitative grading scales - cardiac, diaphragm, muscular, pulmonary, gastric, hepatic and splenic damage score, respectively. Morfological structure of different tissues treated with of 0.1 LD50 of all reactivators were similar to those evaluated in the control groups. Focal and reversible degenerative and vascular changes, were established in tissue samples after treatment with 0.5 LD50 of asoxime, obidoxime and K027 (p < 0.01 vs. control group). Acute alterations were developed in tissue samples within 7 days following treatment with 0.5 LD50 and 1.0 LD50 of all reactivators. The most severe tissue alterations were found in rats treated with .0 LD50 of K048 and K075 (p < 0.001 vs. control and asoxime group, respectively). Our results showed that all AChE reactivators given by a single, high, unitary dose regimen, have adverse effect not only on the main visceral and muscular tissues, but on the whole rat as well, but the exact cause-effect relationship causing cellular injury remains to be established in further investigation. [ABSTRACT FROM AUTHOR]

Progress in the development of biodegradable ionic liquids (ILs) [1] allowed finding sustainable fragments to assist the synthesis of sustainable molecules by means of "benign by design" approach. Based on our recent experience in creating micellar catalytic systems for decomposition of organophosphates [2, 3] we have elaborated the following oxime-functionalized low-toxic biodegradable ILs as potential AcChE reactivators: amide/ester linked (amino acid free) IL (I) as well as L-alanine (II) and L-phenylalanine (III) containing compounds with pyridinium aldoxime moiety in cationic part. Variation of amino acid variation (e.g. Me for I and phenyl for II) can help us to analyze a role of hydrophobicity of IL's cation in AcChE reactivation. The reactivation capacity of novel ILs were evaluated towards AcChE inhibited by typical toxic organophosphate agents. The regularities of antidotal activity of studied compounds are to use in the further improvement of their structures. [ABSTRACT FROM AUTHOR]

K-oximes were developed as modern drug candidates acting as AChE reactivators. In this study, it has been investigated which interspecies and intergender differences changes could be observed in Wistar rats and Swiss mice, both genders, after the treatment with increasing doses of selected acetylcholinesterase reactivators - asoxime, obidoxime, K027, K048, and K075. After the 24 h, a number of died animals was counted and the median lethal dose (LD 50) for each oxime was calculated. By using the intramuscular route of administration, asoxime and K027 had the least toxicity in female rats (640.21 mg/kg and 686.08 mg/kg), and in female mice (565.75 mg/kg and 565.74 mg/kg), respectively. Moreover, asoxime and K027 showed 3, 4 or 8 times less acute toxicity in comparison to K048, obidoxime and K075, respectively. Beyond, K075 had the greatest toxicity in male rats (81.53 mg/kg), and in male mice (57.34 mg/kg), respectively. Our results can help to predict likely adverse toxic effects, target organ systems and possible outcome in the event of massive human overexposure, and in establishing risk categories or in dose selection for the initial repeated dose toxicity tests to be conducted for each oxime. • Asoxime and K027 had the least acute toxicity in female rats, and in female mice. • K075 had the greatest acute toxicity in male rats, and in male mice. • Asoxime and K027 had 3, 4 and 8 times less LD 50 than K048, obidoxime and K075. [ABSTRACT FROM AUTHOR]

Oxidative stress status, as a disruption of redox homeostasis, in the blood sera of Wistar rats caused by repeated application of selected acetylcholinesterase reactivators - asoxime, obidoxime, K027, K048, K074, and K075 were evaluated. Throughout this study, each oxime in a dose of 0.1 of LD 50 /kg im was given 2x/week for 4 weeks. Then, seven days after the last oximes' application, markers of lipid peroxidation (malondialdehyde, MDA), and protein oxidation (advanced oxidation protein products, AOPP), as well as the activity of antioxidant enzymes (catalase, CAT, superoxide dismutase, SOD, reduced glutathione, GSH, and oxidized glutathione, GSSG), were determined. Oxidative stress parameters, MDA and AOPP were significantly highest in the K048-, K074- and K075-treated groups (p < 0.001). The activity of CAT was significantly elevated in the obidoxime-treated group (p < 0.05), while treatment with K027, K048, and K074 induced high elevation in SOD levels (p < 0.01, p < 0.001). Interestingly, the activity of GSH in each oxime-treated group was significantly elevated. Unlike, treatment with obidoxime caused elevation in GSSG levels (p < 0.01). As a continuation of our previously published data, these results assure that applied oximes following subacute treatment ameliorated the oxidative status and further adverse systemic toxic effects in rats. • Treatment by asoxime and K027 reduces markers of lipid peroxidation in rats. • Oxime K048, K074 and K075 elevated the protein oxidation levels in treated rats. • The activity of catalase was markedly elevated in the obidoxime-treated rats. • The superoxide dismutase levels increased in K027-, K048- and K074-treated rats. • Each oxime induced elevation of glutathione peroxidase in treated rats. [ABSTRACT FROM AUTHOR]

Abstract: A toxic effect of highly toxic nervous agents is irreversible inhibition of vitally important enzyme acethylcholinesterase (AChE). Inhibition of AChE results in accumulation of acetylcholine (ACh) at the synaptic cleft of the cholinergic neurons, leading to overstimulation of cholinergic receptors. The highly toxic nature of tabun has been known for many years, but there are still serious limitations to the antidotal therapy. In this paper a bispyridinium compound K027 [1-(4-hydroxyiminomethylpyridinium)-3-(-4-carbamoylpyridinium) propane dibromide] was tested as potential antidote in tabun poisoned mice. Oxime TMB-4 was included for comparison. The therapeutic efficacy of applied antidotal regimens was tested as pretreatment given 15min before tabun poisoning and/or as therapy given 1min after tabun poisoning. Using oxime K027 (25% of its LD50) plus atropine as both, pretreatment and therapy, we showed that this combination can protect mice 8 times better than the therapy alone. Under these experimental conditions we confirmed good antidotal efficacy of K027. Moreover, its low acute toxicity is as much as beneficial effect in contrast to high toxicity of currently used TMB-4. [ABSTRACT FROM AUTHOR]

Abstract: Kinetic parameters were evaluated for inhibition of native and reactivation of tabun-inhibited human erythrocyte acetylcholinesterase (AChE, EC 3.1.1.7) and human plasma butyrylcholinesterase (BChE, EC 3.1.1.8) by three bispyridinium para-aldoximes with butane (K074), but-2-ene (K075) or xylene-like linker (K114). Tested aldoximes reversibly inhibited both cholinesterases with the preference for binding to the native AChE. Both cholinesterases showed the highest affinity for K114 (K i was 0.01mM for AChE and 0.06mM for BChE). The reactivation of tabun-inhibited AChE was efficient by K074 and K075. Their overall reactivation rate constants were around 2000min−1 M−1, which is seven times higher than for the classical bispyridinium para-aldoxime TMB-4. The reactivation of tabun-inhibited AChE assisted by K114 was slow and reached 90% after 20h. Since the aldoxime binding affinity of tabun-inhibited AChE was similar for all tested aldoximes (and corresponded to their K i ), the rate of the nucleophilic displacement of the phosphoryl-moiety from the active site serine was the limiting factor for AChE reactivation. On the other hand, none of the aldoximes displayed a significant reactivation of tabun-inhibited BChE. Even after 20h, the reactivation maximum was 60% for 1mM K074 and K075, and only 20% for 1mM K114. However, lower BChE affinities for K074 and K075 compared to AChE suggest that the fast tabun-inhibited AChE reactivation by these compounds would not be obstructed by their interactions with BChE in vivo. [Copyright &y& Elsevier]

We report a green chemistry approach for preparation of oxime-functionalized ILs as AChE reactivators: amide/ester linked IL, l -alanine, and l -phenylalanine derived salts bearing pyridinium aldoxime moiety. The reactivation capacities of the novel oximes were evaluated towards AChE inhibited by typical toxic organophosphates, sarin (GB), VX, and paraoxon (PON). The studied compounds are mostly non-toxic up to the highest concentrations screened (2 mM) towards Gram-negative and Gram-positive bacteria cell lines and both filamentous fungi and yeasts in the in vitro screening experiments as well as towards the eukaryotic cell (CHO–K1 cell line). Introduction of the oxime moiety in initially biodegradable structure decreases its ability to biodegradation. The compound 3d was shown to reveal remarkable activity against the AChE inhibited by VX, exceeding conventional reactivators 2-PAM and obidoxime. The regularities on antidotal activity, cell viability, plasma stability, biodegradability as well as molecular docking study of the newly synthesized oximes will be used for further improvement of their structures. [Display omitted] • Series of AChE reactivators were synthesized using sustainable IL-based molecular platforms. • Physico-chemical properties and toxicity of novel reactivators were evaluated. • Reactivation capacity were evaluated towards AChE inhibited by typical toxic organophosphates. • Selected oximes demonstrate activity, exceeding conventional reactivators. [ABSTRACT FROM AUTHOR]

Oxidative stress status and morphological injuries in the brain of Wistar rats induced by repeated application of selected acetylcholinesterase reactivators – asoxime, obidoxime, K027, K048, K074, and K075 were evaluated. Each oxime in a dose of 0.1 of LD 50 /kg im was given 2x/week for 4 weeks. Markers of lipid peroxidation (malondialdehyde, MDA), and protein oxidation (advanced oxidation protein products, AOPP), as well as the activity of antioxidant enzymes (catalase, CAT, superoxide dismutase, SOD, glutathione reductase, GR, and glutathione peroxidase, GPx), were estimated in the brain tissue homogenates on day 35 of the study. Brain alterations were carefully quantified by semiquantitative grading scales – brain damage score (BDS). Oxidative stress parameters, MDA and AOPP were significantly highest in the asoxime-, obidoxime- and K075-treated groups (p < 0.001). The activity of SOD and CAT was significantly elevated in the obidoxime-, K048-, and K075-treated groups (p < 0.001). Besides, GR was markedly decreased in the obidoxime- and K074-treated groups (p < 0.01), while treatment with K048, K074 and K075 induced extremely high elevation in GPx levels (p < 0.001). In the same groups of rats, brain alterations associated with polymorphonuclear cell infiltrate were significantly more severe than those observed in animals receiving only asoxime or K027 (p < 0.001). The presented results confirmed that treatment with different oximes significantly improved the oxidative status and attenuated signs of inflammation in rats' brains. Presented results, together with our previously published data can help to predict likely adverse systemic toxic effects, and target organ systems, which are crucial for establishing risk categories, as well as in dose selection of K-oximes as drug candidates. • K-oximes improve the oxidative status of rats' brains following subacute exposure. • Selected K-oximes potently elevate MDA, AOPP, SOD, CAT and GPx levels in rats' brains. • Treatment with K027 extraordinarily attenuates signs of inflammation in rats' brains. [ABSTRACT FROM AUTHOR]

The highly toxic nature of tabun has been known for many years, but there are still serious limitations to antidotal therapy. In this study, we used rats as an experimental model to evaluate the efficiency of bispyridinium para-oxime K203 as therapy against tabun poisoning as well as to examine if induction of oxidative stress is linked to organophosphate toxicity. K203 showed high potency in counteracting tabun poisoning. Either alone or in combination with atropine, this oxime significantly increased cholinesterase activity at 0.5 and 1 h compared to untreated rats poisoned with tabun. Simultaneous measurements of markers of oxidative stress (lipid peroxidation and superoxide dismutase) showed that tabun poisoning, but also therapy (oxime alone or oxime plus atropine) applied immediately after tabun poisoning, could generate free radical species that may cause oxidative stress in rats. [ABSTRACT FROM AUTHOR]

Abstract: Administration of acetylcholinesterase (AChE) reactivators (oximes) is usually used in order to counteract the poisoning effects of nerve agents. The possibility was suggested that oximes may show some therapeutic and/or adverse effects through their action in central nervous system. There are no sufficient data about interaction of oximes with monoaminergic neurotransmitter''s systems in the brain. Oxime-type AChE reactivators pralidoxime, obidoxime, trimedoxime, methoxime and HI-6 were tested for their potential to affect the activity of monoamine oxidase of type A (MAO-A) and type B (MAO-B) in crude mitochondrial fraction of pig brains. The compounds were found to inhibit fully MAO-A with half maximal inhibitory concentration (IC 50) of 0.375mmol/l (pralidoxime), 1.53mmol/l (HI-6), 2.31mmol/l (methoxime), 2.42mmol/l (obidoxime) and 4.98mmol/l (trimedoxime). Activity of MAO-B was fully inhibited by HI-6 and pralidoxime only with IC 50 4.81mmol/l and 11.01mmol/l, respectively. Methoxime, obidoxime and trimedoxime displayed non-monotonic concentration dependent effect on MAO-B activity. Because oximes concentrations effective for MAO inhibition could not be achieved in vivo at the cerebral level, we suppose that oximes investigated do not interfere with brain MAO at therapeutically relevant concentrations. [Copyright &y& Elsevier]

Abstract: In this work a theoretical methodology for evaluation of the association and kinetic reactivation constants of oximes using the Molegro® and Spartan® softwares was proposed and validated facing in vitro data previously reported in the literature. Results showed a very good agreement between the theoretical binding free energies of the reactivators and experimental data, suggesting that the proposed methodology could work well in the prediction of kinetic and thermodynamics parameters for oximes that might be helpful for the design and selection of new and more effective oximes. [ABSTRACT FROM AUTHOR]

Abstract: Butyrylcholinesterase is considered to be an endogenous stoichiometric bioscavenger of organophosphorus compounds (OPs), but due to limited concentration of BChE in the organism, stoichiometric reduction of OP is not always sufficient. This can be improved by creating a pseudo-catalytic scavenger adding oximes as reactivators of inhibited exogenous BChE. In order to improve the BChE bioscavenging function in tabun or paraoxon poisoning, we tested in vitro reactivation of phosphorylated human plasma BChE by bispyridinium oximes varying in the length and type of the linker between rings, and in the position of the oxime group on the ring. Among the tested oximes, the most potent reactivators of tabun-inhibited BChE were K117 [1,1′-(2,2′-oxybis(ethane-2,1-diyl))bis(4-hydroxyiminomethyl pyridinium) bromide] and K127 [4-carbamoyl-1-(2-(2-(4-(hydroxyiminomethyl) pyridinium-1-yl)ethoxy)ethyl)pyridinium bromide]. Reactivation by these oximes (1mM) reached about 50% of control activity after only 20min; however, reactivation stopped at 70%. Reactivation of paraoxon-inhibited BChE by all of the selected oximes was slow. Using molecular mechanics, we performed docking of the oximes to tabun-inhibited BChE in order to discuss possible structural modifications of bispyridinium oximes to improve reactivation of phosphorylated BChE. [ABSTRACT FROM AUTHOR]

Abstract: Improving the efficacy of antidotal treatment of poisonings with nerve agents is still a challenge for the scientific community. This study investigated the interactions of four bispyridinium oximes with human erythrocyte acetylcholinesterase (AChE) and their effects on soman- and tabun-poisoned mice. Oximes HI-6 and TMB-4 were used for comparison. These oximes inhibited AchE with inhibitory potency (IC50) ranging from 0.02 to 1.0mM. The best reactivating potency (%R) was obtained with K074, when AChE was inhibited by tabun. The protective potency (P50) of all oximes in human erythrocyte AChE inhibited by soman and tabun could not be determined. In tabun-poisoned mice very good antidotal efficacy was obtained with K027, K048, and K074, which makes them interesting for future investigation. The combination of HI-6 and atropine is the therapy of choice for soman poisoning. [Copyright &y& Elsevier]

Abstract: Oximes K033 [1,4-bis(2-hydroxyiminomethylpyridinium) butane dibromide] and K048 [1-(4-hydroxyiminomethylpyridinium)-4-(4-carbamoylpyridinium) butane dibromide] were tested as pretreatment drugs in tabun-poisoned mice followed by treatment with atropine plus K033, K048, K027 [1-(4-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium) propane dibromide], TMB-4 [1,3-bis(4-hydroxyiminomethylpyridinium) propane dibromide] and HI-6 [(1-(2-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium)-2-oxapropane dichloride)]. Oxime doses of 25% or 5% of its LD50 were used for pretreatment 15min before tabun-poisoning and for treatment 1min after tabun administration to mice. The best therapeutic effect was obtained when oxime K048 (25% of its LD50) was used in both pretreatment and treatment with atropine. This regiment insured survival of all tested animals after the application of 10 LD50 of tabun. In addition, since butyrylcholinesterase (BChE; EC 3.1.1.8) is considered an endogenous bioscavenger of anticholinesterase compounds and its interactions with oximes could be masked by AChE interactions, we evaluated kinetic parameters for interactions of tested oximes with native and tabun-inhibited human plasma BChE and compared them with results obtained previously for human erythrocyte acetylcholinesterase (AChE; EC 3.1.1.7). Progressive inhibition of BChE by tabun was slightly faster than that of AChE. The reactivation of tabun-inhibited BChE by oximes was very slow, and BChE binding affinity for oximes was lower than AChE''s. Therefore, BChE could scavenge tabun prior to AChE inhibition, but fast oxime-assisted reactivation of tabun-inhibited AChE or protection of AChE by oxime against inhibition with tabun would not be obstructed by interaction between BChE and oximes. [Copyright &y& Elsevier]

Abstract: The increased concern about terrorist use of nerve agents prompted us to search for new more effective oximes against tabun and soman poisoning. We investigated the interactions of five bispyridinium oximes: K027 [1-(4-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium) propane dibromide], K048 [1-(4-hydroxyiminomethylpyridinium)-4-(4-carbamoylpyridinium) butane dibromide], K033 [1,4-bis(2-hydroxyiminomethylpyridinium) butane dibromide], TMB-4 [1,3-bis(4-hydroxyiminomethylpyridinium) propane dibromide] and HI-6 [(1-(2-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium)-2-oxapropane dichloride)] with human erythrocyte acetylcholinesterase (AChE; E.C. 3.1.1.7) and their effects on tabun- and soman-poisoned mice. All the oximes reversibly inhibited AChE, and the enzyme-oxime dissociation constants were between 17 and 180μM. Tabun-inhibited AChE was completely reactivated by TMB-4, K027 and K048, with the overall reactivation rate constants of 306, 376 and 673min−1 M−1, respectively. The reactivation of tabun-inhibited AChE by K033 reached 50% after 24h, while HI-6 failed to reactivate any AChE at all. Soman-inhibited AChE was resistant to reactivation by 1mM oximes. All studied oximes protected AChE from phosphorylation with both soman and tabun. In vivo experiments showed that the studied oximes were relatively toxic to mice; K033 was the most toxic (LD50 =33.4mg/kg), while K027 was the least toxic (LD50 =672.8mg/kg). The best antidotal efficacy was obtained with K048, K027 and TMB-4 for tabun poisoning, and HI-6 for soman poisoning. Moreover, all tested oximes showed no cytotoxic effect on several cell lines in concentrations up to 0.8mM. The potency of the oximes K048 and K027 to protect mice from five-fold LD50 of tabun and their low toxicity make these compounds leading in the therapy of tabun poisoning. The combination of HI-6 and atropine is the therapy of choice for soman poisoning. [Copyright &y& Elsevier]