Mono-quaternary pyridinium oximes derived from K-oximes K027, K048 and K203 were designed, synthesized and evaluated for the reactivation of organophosphate-inhibited cholinesterases. The incorporation of the halogen atoms to the structure decreased the pKa value of the oxime group resulting in an increased formation of oximate necessary for reactivation. The stability and pKa values were found to be similar to analogous bis-quaternary compounds. Some mono-quaternary oximes resulted as relatively strong inhibitors of human acetylcholinesterase. Nevertheless, the reactivation ability of mono-quaternary oximes for organophosphate-inhibited cholinesterases was lower compared to their bis-quaternary analogues. These results were further confirmed by the determination of reactivation kinetics, when in some cases novel compounds showed improvement reactivation compared to the tested standards, but no improvement to bis-quaternary K-oximes. A computational study investigated reactivation process for K027, and its two analogues for VX-inhibited AChE. This study revealed slight differences between reactivation of mono-quaternary and bis-quaternary oximes. Abbreviations: 2-PAM, pralidoxime; AChE, acetylcholinesterase; ACN, acetonitrile; ATCI, acetylcholine iodide; BChE, butyrylcholinesterase; BTCI, butyrylcholine iodide; Bu3SnSnBu3, bis(tributyltin) Et2O, diethyl ether; ChEs, cholinesterases; CNS, central nervous system; DAD, diode array detector; DIBAL-H, diisobutylaluminium hydride; DMF, dimethylformamide; DMSO, dimethyl sulfoxide; DTNB, 5,5́-dithiobis-2-nitrobenzoic acid; Et3N, triethylamine; EtOAc, ethyl acetate; EWG, electron withdrawing group; HI-6, asoxime; hrAChE, human recombinant acetylcholinesterase; hrBChE, human recombinant butyrylcholinesterase; hrChEs, human recombinant cholinesterases; HPLC, high-performance liquid chromatography; HRMS, high-resolution mass spectrometry; KD, dissociation constant; kr, first-order reactivation rate constant; kr2, second-order reactivation rate constant; LüH-6, obidoxime; MeOH, methanol; MM, molecular mechanics; MMC-4, methoxime; m.p., melting point; NCIs, non-covalent interactions; NEDPA, 4-nitrophenyl ethyl dimethylphosphoramidate; NEMP, 4-nitrophenyl ethyl methylphosphonate; NIMP, isopropyl methylphosphonate; NMR, nuclear magnetic resonance spectroscopy; OPs, organophosphates; PBS, phosphate-buffered saline; Pd(dppf)Cl2.CH2Cl2, [1,1'-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) in complex with dichloromethane; pKa, negative decimal logarithm of the dissociation constant; POX, paraoxon; PPh3, triphenylphosphine; QM, quantum mechanics; rt, room temperature; SN2, bimolecular nucleophilic substitution; SNAc, nucleophilic acyl substitution; THF, tetrahydrofuran; TMC-4, trimedoxime; TNB, 5-thio-2-nitrobenzoic acid; UHPLC, ultra high-performance liquid chromatography; UV, ultraviolet; UV-VIS, ultraviolet-visible.
- Klíčová slova
- Cholinesterase, Inhibition, Organophosphate, Oxime, Reactivation,
- MeSH
- acetylcholinesterasa * metabolismus MeSH
- butyrylcholinesterasa metabolismus MeSH
- cholinesterasové inhibitory * farmakologie chemie chemická syntéza MeSH
- halogenace MeSH
- kinetika MeSH
- lidé MeSH
- molekulární struktura MeSH
- oximy * chemie farmakologie chemická syntéza MeSH
- pyridinové sloučeniny chemie farmakologie chemická syntéza MeSH
- reaktivátory cholinesterasy farmakologie chemie chemická syntéza MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- acetylcholinesterasa * MeSH
- butyrylcholinesterasa MeSH
- cholinesterasové inhibitory * MeSH
- oximy * MeSH
- pyridinové sloučeniny MeSH
- reaktivátory cholinesterasy MeSH
Six novel brominated bis-pyridinium oximes were designed and synthesized to increase their nucleophilicity and reactivation ability of phosphorylated acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Their pKa was valuably found lower to parent non-halogenated oximes. Stability tests showed that novel brominated oximes were stable in water, but the stability of di-brominated oximes was decreased in buffer solution and their degradation products were prepared and characterized. The reactivation screening of brominated oximes was tested on AChE and BChE inhibited by organophosphorus surrogates. Two mono-brominated oximes reactivated AChE comparably to non-halogenated analogues, which was further confirmed by reactivation kinetics. The acute toxicity of two selected brominated oximes was similar to commercially available oxime reactivators and the most promising brominated oxime was tested in vivo on sarin- and VX-poisoned rats. This brominated oxime showed interesting CNS distribution and significant reactivation effectiveness in blood. The same oxime resulted with the best protective index for VX-poisoned rats.
- Klíčová slova
- Cholinesterase, Nerve agent, Nucleophile, Organophosphate, Oxime, Reactivation,
- MeSH
- acetylcholinesterasa * metabolismus účinky léků MeSH
- butyrylcholinesterasa * metabolismus MeSH
- chemické bojové látky toxicita MeSH
- cholinesterasové inhibitory * toxicita farmakologie MeSH
- halogenace MeSH
- krysa rodu Rattus MeSH
- nervová bojová látka * toxicita MeSH
- organothiofosforové sloučeniny * toxicita MeSH
- oximy * farmakologie chemie MeSH
- potkani Wistar MeSH
- pyridinové sloučeniny farmakologie MeSH
- reaktivátory cholinesterasy * farmakologie chemie MeSH
- sarin * toxicita MeSH
- stabilita léku MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- acetylcholinesterasa * MeSH
- butyrylcholinesterasa * MeSH
- chemické bojové látky MeSH
- cholinesterasové inhibitory * MeSH
- nervová bojová látka * MeSH
- organothiofosforové sloučeniny * MeSH
- oximy * MeSH
- pyridinové sloučeniny MeSH
- reaktivátory cholinesterasy * MeSH
- sarin * MeSH
- VX MeSH Prohlížeč
The fluorinated bis-pyridinium oximes were designed and synthesized with the aim of increasing their nucleophilicity and potential to reactivate phosphorylated human recombinant acetylcholinesterase (AChE) and human purified plasmatic butyrylcholinesterase (BChE) in relation to chlorinated and non-halogenated oxime analogues. Compared to non-halogenated oximes, halogenated oximes showed lower pKa of the oxime group (fluorinated < chlorinated < non-halogenated) along with higher level of oximate anion formation at the physiological pH, and had a higher binding affinity of both AChE and BChE. The stability tests showed that the fluorinated oximes were stable in water, while in buffered environment di-fluorinated oximes were prone to rapid degradation, which was reflected in their lower reactivation ability. Mono-fluorinated oximes showed comparable reactivation to non-halogenated (except asoxime) and mono-chlorinated oximes in case of AChE inhibited by sarin, cyclosarin, VX, and tabun, but were less efficient than di-chlorinated ones. The same trend was observed in the reactivation of inhibited BChE. The advantage of halogen substituents in the stabilization of oxime in a position optimal for in-line nucleophilic attack were confirmed by extensive molecular modelling of pre-reactivation complexes between the analogue oximes and phosphorylated AChE and BChE. Halogen substitution was shown to provide oximes with additional beneficial properties, e.g., fluorinated oximes gained antioxidative capacity, and moreover, halogens themselves did not increase cytotoxicity of oximes. Finally, the in vivo administration of highly efficient reactivator and the most promising analogue, 3,5-di-chloro-bispyridinium oxime with trimethylene linker, provided significant protection of mice exposed to sarin and cyclosarin.
- Klíčová slova
- Cholinesterase, Nerve agent, Organophosphate, Oxime, Reactivation,
- MeSH
- acetylcholinesterasa metabolismus MeSH
- butyrylcholinesterasa metabolismus MeSH
- cholinesterasové inhibitory chemie MeSH
- halogeny MeSH
- myši MeSH
- nervová bojová látka * farmakologie MeSH
- organofosforové sloučeniny MeSH
- oximy chemie MeSH
- reaktivátory cholinesterasy * chemie MeSH
- sarin chemie MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- acetylcholinesterasa MeSH
- butyrylcholinesterasa MeSH
- cholinesterasové inhibitory MeSH
- halogeny MeSH
- nervová bojová látka * MeSH
- organofosforové sloučeniny MeSH
- oximy MeSH
- reaktivátory cholinesterasy * MeSH
- sarin MeSH
