The current study imposes a new class of organophosphorus (OP)-inhibited cholinesterase reactivators by conceptualizing a family of asymmetric bisoximes with various reactivating scaffolds. Several novel nucleophilic warheads were investigated, putting forward 29 novel reactivating options, by evaluating their nucleophilicity and ability to directly decompose OP compounds. Adopting the so-called zwitterionic strategy, 17 mono-oxime and nine bisoxime reactivators were discovered with major emphasis on the bifunctional-moiety approach. Compounds were compared with clinically used standards and other known experimentally highlighted reactivators. Our results clearly favor the concept of asymmetric bisoximes as leading reactivators in terms of efficacy and versatility. These top-ranked compounds were characterized in detail by reactivation kinetics parameters and evaluated for potential CNS availability. The highlighted molecules 55, 57, and 58 with various reactivating warheads, surpassed the reactivating potency of pralidoxime and several notable uncharged reactivators. The versatility of lead drug candidate 55 was also inspected on OP-inhibited butyrylcholinesterase, revealing a much higher rate compared to existing clinical antidotes.
- MeSH
- acetylcholinesterasa metabolismus MeSH
- antidota chemie farmakologie MeSH
- butyrylcholinesterasa * metabolismus chemie MeSH
- cholinesterasové inhibitory chemie farmakologie MeSH
- kinetika MeSH
- lidé MeSH
- organofosforové sloučeniny chemie MeSH
- otrava organofosfáty * farmakoterapie MeSH
- oximy * chemie farmakologie MeSH
- reaktivátory cholinesterázy * chemie farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
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.
- MeSH
- acetylcholinesterasa MeSH
- antidota farmakologie MeSH
- butyrylcholinesterasa MeSH
- cholinesterasové inhibitory toxicita MeSH
- fosfor MeSH
- krysa rodu rattus MeSH
- kyslík MeSH
- lidé MeSH
- oximy farmakologie MeSH
- pralidoximové sloučeniny * MeSH
- pyridinové sloučeniny farmakologie MeSH
- reaktivátory cholinesterázy * farmakologie MeSH
- taurin analogy a deriváty MeSH
- trimedoxim farmakologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Oxime reactivators of acetylcholinesterase (AChE) are used as causal antidotes for intended and unintended poisoning by organophosphate nerve agents and pesticides. Despite all efforts to develop new AChE reactivators, none of these drug candidates replaced conventional clinically used oximes. In addition to the therapeutic efficacy, determining the safety profile is crucial in preclinical drug evaluation. The exact mechanism of oxime toxicity and the structure-toxicity relationship are subjects of ongoing research, with oxidative stress proposed as a possible mechanism. In the present study, we investigated four promising bispyridinium oxime AChE reactivators, K048, K074, K075, and K203, and their ability to induce oxidative stress in vitro. Cultured human hepatoma cells were exposed to oximes at concentrations corresponding to their IC50 values determined by the MTT assay after 24 h. Their potency to generate reactive oxygen species, interfere with the thiol antioxidant system, and induce lipid peroxidation was evaluated at 1, 4, and 24 h of exposure. Reactivators without a double bond in the four-carbon linker, K048 and K074, showed a greater potential to induce oxidative stress compared with K075 and K203, which contain a double bond. Unlike oximes with a three-carbon-long linker, the number of aldoxime groups attached to the pyridinium moieties does not determine the oxidative stress induction for K048, K074, K075, and K203 oximes. In conclusion, our results emphasize that the structure of oximes plays a critical role in inducing oxidative stress, and this relationship does not correlate with their cytotoxicity expressed as the IC50 value. However, it is important to note that oxidative stress cannot be disregarded as a potential contributor to the side effects associated with oximes.
- MeSH
- acetylcholinesterasa metabolismus MeSH
- antidota farmakologie MeSH
- buňky Hep G2 MeSH
- cholinesterasové inhibitory toxicita MeSH
- lidé MeSH
- organofosfáty toxicita MeSH
- oxidační stres MeSH
- oximy farmakologie chemie MeSH
- pyridinové sloučeniny farmakologie chemie MeSH
- reaktivátory cholinesterázy * farmakologie chemie MeSH
- uhlík MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
The problem of the efficient treatment of acute organophosphorus (OP) poisoning needs more efforts in the development of a versatile antidote, applicable for treatment of the injuries of both peripheral and central nervous systems. A series of N-H, N-methyl, N-butyl, and N-phenyl derivatives of benzhydroxamic (1a-1d), 3-methoxybenzhydroxamic (2a-2d), 4-methoxybenzhydroxamic (3a-3d) acids, and corresponding salycilhydroxamates (4a-4d) was prepared. Their predicted hydrophobicity (log P) was evaluated as regards to ВВВ score by the open access cheminformatics tools; prediction of the passive transport across the BBB was found by means on the parallel artificial membrane permeability assay (PAMPA). The data on reactivation capacity of human acetylcholinesterase (HssAChE) inhibited by GB, VX, and paraoxon was supported by molecular docking study on binding to the active site of the AChE, viability study against mammalian cells (Chinese hamster ovary CHO-K1), and biodegradability (Closed Bottle test OECD 301D). Among the studied compounds, N-butyl derivatives have better balanced combination of properties; among them, N-butylsalicylhydroxamic acid is most promising. The studied compounds demonstrate modest reactivation capacity; change of N-H by N-Me ensures the reactivation capacity in studied concentrations on all studied OP substrates; among N-butyl derivatives, the N-butylsalicylhydroxamic acid demonstrates most promising results within the series. The found regularities may lead to selection of perspective structures to complement current formulations for medical countermeasures against poisoning by organophosphorus toxicants.
- MeSH
- acetylcholinesterasa metabolismus MeSH
- antidota farmakologie MeSH
- CHO buňky MeSH
- cholinesterasové inhibitory chemie farmakologie MeSH
- Cricetulus MeSH
- křečci praví MeSH
- lidé MeSH
- otrava organofosfáty * MeSH
- oximy chemie MeSH
- reaktivátory cholinesterázy * chemie farmakologie MeSH
- simulace molekulového dockingu MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- křečci praví MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
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.
- MeSH
- acetylcholinesterasa MeSH
- antidota farmakologie MeSH
- cholinesterasové inhibitory toxicita MeSH
- lidé MeSH
- nervová bojová látka * toxicita MeSH
- oximy farmakologie MeSH
- reaktivátory cholinesterázy * farmakologie MeSH
- simulace molekulového dockingu MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
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.
- MeSH
- acetylcholinesterasa metabolismus MeSH
- antidota farmakologie MeSH
- cholinesterasové inhibitory chemie toxicita MeSH
- lidé MeSH
- nervová bojová látka * toxicita MeSH
- organofosfáty MeSH
- oximy chemie farmakologie MeSH
- pyridinové sloučeniny farmakologie MeSH
- reaktivátory cholinesterázy * farmakologie MeSH
- simulace molekulového dockingu MeSH
- trimedoxim farmakologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
AIM: The comparison of neuroprotective and central reactivating effects of the oxime K870 in combination with atropine with the efficacy of standard antidotal treatment in tabun-poisoned rats. METHODS: The neuroprotective effects of antidotal treatment were determined in rats poisoned with tabun at a sublethal dose using a functional observational battery 2 h and 24 h after tabun administration, the tabun-induced brain damage was investigated by the histopathological evaluation and central reactivating effects of oximes was evaluated by the determination of acetylcholinesterase activity in the brain using a standard spectrophotometric method. RESULTS: The central reactivating efficacy of a newly developed oxime K870 roughly corresponds to the central reactivating efficacy of pralidoxime while the ability of the oxime HI-6 to reactivate tabun-inhibited acetylcholinesterase in the brain was negligible. The ability of the oxime K870 to decrease tabun-induced acute neurotoxicity was slightly higher than that of pralidoxime and similar to the oxime HI-6. These results roughly correspond to the histopathological evaluation of tabun-induced brain damage. CONCLUSION: The newly synthesized oxime K870 is not a suitable replacement for commonly used oximes in the antidotal treatment of acute tabun poisonings because its neuroprotective efficacy is only slightly higher or similar compared to studied currently used oximes.
- MeSH
- acetylcholinesterasa MeSH
- antidota farmakologie MeSH
- chemické bojové látky * toxicita MeSH
- cholinesterasové inhibitory farmakologie MeSH
- jedy * MeSH
- krysa rodu rattus MeSH
- organofosfáty * MeSH
- oximy * farmakologie MeSH
- potkani Wistar MeSH
- pralidoximové sloučeniny MeSH
- pyridinové sloučeniny * farmakologie MeSH
- reaktivátory cholinesterázy * farmakologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
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.
BACKGROUND: Neurotoxic chemical warfare agents can be classified as some of the most dangerous chemicals for humanity. The most effective of those agents are the Organophosphates (OPs) capable of restricting the enzyme Acetylcholinesterase (AChE), which in turn, controls the nerve impulse transmission. When AChE is inhibited by OPs, its reactivation can be usually performed through cationic oximes. However, until today, it has not been developed one universal defense agent, with complete effective reactivation activity for AChE inhibited by any of the many types of existing neurotoxic OPs. For this reason, before treating people intoxicated by an OP, it is necessary to determine the neurotoxic compound that was used for contamination, in order to select the most effective oxime. Unfortunately, this task usually requires a relatively long time, raising the possibility of death. Cationic oximes also display a limited capacity of permeating the Blood-Brain Barrier (BBB). This fact compromises their capacity to reactivating AChE inside the nervous system. METHODS: We performed a comprehensive search on the data about OPs available on the scientific literature today in order to cover all the main drawbacks still faced in the research for the development of effective antidotes against those compounds. RESULTS: Therefore, this review about neurotoxic OPs and the reactivation of AChE, provides insights for the new agents' development. The most expected defense agent is a molecule without toxicity and effective to reactivate AChE inhibited by all neurotoxic OPs. CONCLUSION: To develop these new agents, the application of diverse scientific areas of research, especially theoretical procedures as computational science (computer simulation, docking and dynamics), organic synthesis, spectroscopic methodologies, biology, biochemical and biophysical information, medicinal chemistry, pharmacology and toxicology, is necessary.
To date, the only treatments developed for poisoning by organophosphorus compounds, the most toxic chemical weapons of mass destruction, have exhibited limited efficacy and versatility. The available causal antidotes are based on reactivation of the enzyme acetylcholinesterase (AChE), which is rapidly and pseudo-irreversibly inhibited by these agents. In this study, we developed a novel series of monoquaternary reactivators combining permanently charged moieties tethered to position 6- of 3-hydroxypyridine-2-aldoxime reactivating subunit. Highlighted representatives (21, 24, and 27; also coded as K1371, K1374, and K1375, respectively) that contained 1-phenylisoquinolinium, 7-amino-1-phenylisoquinolinium and 4-carbamoylpyridinium moieties as peripheral anionic site ligands, respectively, showed efficacy superior or comparable to that of the clinically used standards. More importantly, these reactivators exhibited wide-spectrum efficacy and were minutely investigated via determination of their reactivation kinetics in parallel with molecular dynamics simulations to study their mechanisms of reactivation of the tabun-inhibited AChE conjugate. To further confirm the potential applicability of these candidates, a mouse in vivo assay was conducted. While K1375 had the lowest acute toxicity and the most suitable pharmacokinetic profile, the oxime K1374 with delayed elimination half-life was the most effective in ameliorating the signs of tabun toxicity. Moreover, both in vitro and in vivo, the versatility of the agents was substantially superior to that of clinically used standards. Their high efficacy and broad-spectrum capability make K1374 and K1375 promising candidates that should be further investigated for their potential as nerve agents and insecticide antidotes.
- MeSH
- acetylcholinesterasa účinky léků metabolismus MeSH
- antidota chemická syntéza chemie farmakologie MeSH
- myši inbrední BALB C MeSH
- myši MeSH
- oximy chemická syntéza chemie farmakologie MeSH
- reaktivátory cholinesterázy chemická syntéza chemie farmakologie MeSH
- simulace molekulární dynamiky MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
