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
We report for the first time the efficient use of accelerated solvent extraction (ASE) for extraction of ricin to analytical purposes, followed by the combined use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and MALDI-TOF MS/MS method. That has provided a fast and unambiguous method of ricin identification for in real cases of forensic investigation of suspected samples. Additionally, MALDI-TOF MS was applied to characterize the presence and the toxic activity of ricin in irradiated samples. Samples containing ricin were subjected to ASE, irradiated with different dosages of gamma radiation, and analyzed by MALDI-TOF MS/MS for verification of the intact protein signal. For identification purposes, samples were previously subjected to SDS-PAGE, for purification and separation of the chains, followed by digestion with trypsin, and analysis by MALDI-TOF MS/MS. The results were confirmed by verification of the amino acid sequences of some selected peptides by MALDI-TOF MS/MS. The samples residual toxic activity was evaluated through incubation with a DNA substrate, to simulate the attack by ricin, followed by MALDI-TOF MS/MS analyses.
- MeSH
- aceton chemie MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- hexany chemie MeSH
- peptidy analýza chemie MeSH
- ricin analýza chemie MeSH
- rozpouštědla chemie MeSH
- sekvence aminokyselin MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- tandemová hmotnostní spektrometrie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
