The aim of this study was to determine the antidotal potential of the chlorinated oxime K870 compared to obidoxime, as a monotherapy and in combination with atropine, in paraoxon (POX)-poisoned rats. The treatment doses of oximes were chosen as 20% of their LD50 values. The protective ratio (PR) of oxime K870 with atropine was significantly higher than that of obidoxime with atropine (68.8 and 125.0, respectively). In the biochemical part of the experiment POX subcutaneously (s.c.) (0.75% LD50) was administered and followed by oxime K870 or obidoxime i.m. 1 min later. Acetylcholinesterase (AChE) activity was determined spectrophotometrically in cerebrum, cerebellum, brainstem, diaphragm, and erythrocytes. Carboxylesterase activity was determined in plasma and liver. Both oximes successfully reactivated AChE in brain (cerebrum, cerebellum, and brainstem), diaphragm and erythrocytes, but the oxime K870 performed better than obidoxime. Both oximes reactivated carboxylesterase, obidoxime better in plasma and oxime K870 better in liver. In conclusion, the oxime K870, when co-administered with atropine, is a more effective antidote than the obidoxime-atropine combination in POX-poisoned rats.
- Klíčová slova
- Cholinesterase, K870, obidoxime, organophosphate, paraoxon, reactivator,
- Publikační typ
- časopisecké články MeSH
Organophosphorus compounds are highly toxic irreversible inhibitors of cholinesterases, causing the disruption of cholinergic functions. Treatment of poisoning includes causal antidotes (oximes) used as reactivators of inhibited cholinesterases, such as pralidoxime. In this work, new halogenated oxime reactivators derived from pralidoxime were developed. The oximes were designed with a halogen substituent that lowers the pK a and enhances oximate formation. Their synthesis, stability, physicochemical properties, inhibition of native cholinesterases, and in vitro reactivation of organophosphate-inhibited cholinesterases were investigated. A series of C4 and C6 halogenated oximes showed instability and their degradation products were identified, while C3 and C5 oximes exhibited sufficient stability for the evaluation. C3 oximes displayed overall low inhibition of cholinesterases and high reactivation ability of organophosphate-inhibited cholinesterases compared to pralidoxime, indicating the significant impact of halogen substitution on reactivation ability.
- Publikační typ
- časopisecké články MeSH
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 cholinesterázy 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 cholinesterázy MeSH
Plasmonic photothermal therapy (PPTT) employing plasmonic gold nanorods (GNRs) presents a potent strategy for eradication of tumors including aggressive brain gliomas. Despite its promise, there is a pressing need for a more comprehensive evaluation of PPTT using sophisticated in vitro models that closely resemble tumor tissues, thereby facilitating the elucidation of therapeutic mechanisms. In this study, we exposed 3D glioma spheroids (tumoroids) to (16-mercaptohexadecyl)trimethylammonium bromide-functionalized gold nanorods (MTAB-GNRs) and a near-infrared (NIR) laser. We demonstrate that the photothermal effect can be fine-tuned by adjusting the nanoparticle concentration and laser power. Depending on the selected parameters, the laser can trigger either regulated or non-regulated cell death (necrosis) in both mouse GL261 and human U-87 MG glioma cell lines, accompanied by translocation of phosphatidylserine in the membrane. Our investigation into the mechanism of regulated cell death induced by PPTT revealed an absence of markers associated with classical apoptosis pathways, such as cleaved caspase 3. Instead, we observed the presence of cleaved caspase 1, gasdermin D, and elevated levels of NLRP3 in NIR-irradiated tumoroids, indicating the activation of pyroptosis. This finding correlates with previous observations of lysosomal accumulation of MTAB-GNRs and the known lysosomal pathway of pyroptosis activation. We further confirmed the absence of toxic breakdown products of GNRs using electron microscopy, which showed no melting or fragmentation of gold nanoparticles under the conditions causing regulated cell death. In conclusion, PPTT using coated gold nanorods offers significant potential for glioma cell elimination occurring through the activation of pyroptosis rather than classical apoptosis pathways.
- Klíčová slova
- Cell death, Glioblastoma, Gold nanorods, Plasmonic photothermal effect, Pyroptosis, Tumor spheroids,
- MeSH
- buněčné sféroidy účinky léků patologie MeSH
- fototermální terapie MeSH
- gliom * patologie farmakoterapie metabolismus MeSH
- kationty chemie farmakologie MeSH
- kovové nanočástice chemie MeSH
- kvartérní amoniové sloučeniny chemie farmakologie MeSH
- lidé MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- nádorové buňky kultivované MeSH
- nanotrubičky * chemie MeSH
- pyroptóza * účinky léků MeSH
- viabilita buněk účinky léků MeSH
- zlato * chemie farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- kationty MeSH
- kvartérní amoniové sloučeniny MeSH
- zlato * MeSH
Cyclophilin D (CypD) is a mitochondrial enzyme widely accepted as a regulator of the mitochondrial permeability transition pore (mPTP). Excessive opening of mPTP is associated with mitochondrial dysfunction and the development of various diseases; thus, suppression of mPTP opening through CypD inhibition presents a promising therapeutic approach. However, only a limited number of selective CypD inhibitors are currently available. In this study, 10 derivatives of 2-(benzyloxy)arylurea similar or identical to previously published CypD/mPTP inhibitors were synthesized. Unlike the original reports that assessed the opening of mPTP at the cellular level, the compounds were tested directly on the purified CypD enzyme to validate their putative mechanism of action. Additionally, the effect of the selected compounds was tested on isolated mitochondria. The obtained results show that the compounds are only weak inhibitors of CypD and mPTP opening, which is in contrast to previous conclusions drawn from the unspecific cellular JC-1 assay.
- Publikační typ
- časopisecké články MeSH
Small molecules that exhibit broad-spectrum enteroviral inhibitory activity by targeting viral replication proteins are highly desired in antiviral drug discovery studies. To discover new human rhinovirus (hRV) inhibitors, we performed a high-throughput screening of 100,000 compounds from the Korea Chemical Bank library. This search led to identification of two phosphatidylinositol-4-kinase IIIβ (PI4KIIIβ) inhibitors having the pyrazolo-pyrimidine core structure, which display moderate anti-rhinoviral activity along with mild cytotoxicity. The results of a study aimed at optimizing the activity of the hit compounds showed that the pyrazolo-pyrimidine derivative 6f exhibits the highest activity (EC50 = 0.044, 0.066, and 0.083 μM for hRV-B14, hRV-A16, and hRV-A21, respectively) and moderate toxicity (CC50 = 31.38 μM). Furthermore, 6f has broad-spectrum activities against various hRVs, coxsackieviruses and other enteroviruses, such as EV-A71, EV-D68. An assessment of kinase inhibition potencies demonstrated that 6f possesses a high and selective kinase inhibition activity against PI4KIIIβ (IC50 value of 0.057 μM) and not against PI4KIIIα (>10 μM). Moreover, 6f exhibits modest hepatic stability (46.9 and 55.3 % remaining after 30 min in mouse and human liver microsomes, respectively). Finally, an in vivo study demonstrated that 6f possesses a desirable pharmacokinetic profile reflected in low systemic clearance (0.48 L∙h-1 kg-1) and modest oral bioavailability (52.4 %). Hence, 6f (KR-26549) appears to be an ideal lead for the development of new antiviral drugs.
- Klíčová slova
- Antiviral compound, Enterovirus, Human rhinovirus, PI4KIIIβ, Pyrazolo-pyrimidine,
- MeSH
- antivirové látky * farmakologie chemie chemická syntéza MeSH
- fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
- inhibitory proteinkinas farmakologie chemie chemická syntéza MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- molekulární struktura MeSH
- myši MeSH
- pyrazoly farmakologie chemie chemická syntéza MeSH
- pyrimidiny * farmakologie chemie chemická syntéza MeSH
- replikace viru * účinky léků MeSH
- Rhinovirus * účinky léků MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antivirové látky * MeSH
- fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
- inhibitory proteinkinas MeSH
- phosphatidylinositol 4-kinase IIIbeta, human MeSH Prohlížeč
- pyrazoly MeSH
- pyrimidiny * MeSH
6-Nitrobenzo[b]thiophene 1,1-dioxide (Stattic) is a potent signal transducer and activator of the transcription 3 (STAT3) inhibitor developed originally for anticancer therapy. However, Stattic harbors several STAT3 inhibition-independent biological effects. To improve the properties of Stattic, we prepared a series of analogues derived from 6-aminobenzo[b]thiophene 1,1-dioxide, a compound directly obtained from the reduction of Stattic, that includes a methoxybenzylamino derivative (K2071) with optimized physicochemical characteristics, including the ability to cross the blood-brain barrier. Besides inhibiting the interleukin-6-stimulated activity of STAT3 mediated by tyrosine 705 phosphorylation, K2071 also showed cytotoxicity against a set of human glioblastoma-derived cell lines. In contrast to the core compound, a part of K2071 cytotoxicity reflected a STAT3 inhibition-independent block of mitotic progression in the prophase, affecting mitotic spindle formation, indicating that K2071 also acts as a mitotic poison. Compared to Stattic, K2071 was significantly less thiol-reactive. In addition, K2071 affected cell migration, suppressed cell proliferation in tumor spheroids, exerted cytotoxicity for glioblastoma temozolomide-induced senescent cells, and inhibited the secretion of the proinflammatory cytokine monocyte chemoattractant protein 1 (MCP-1) in senescent cells. Importantly, K2071 was well tolerated in mice, lacking manifestations of acute toxicity. The structure-activity relationship analysis of the K2071 molecule revealed the necessity of the para-substituted methoxyphenyl motif for antimitotic but not overall cytotoxic activity of its derivatives. Altogether, these results indicate that compound K2071 is a novel Stattic-derived STAT3 inhibitor and a mitotic poison with anticancer and senotherapeutic properties that is effective on glioblastoma cells and may be further developed as an agent for glioblastoma therapy.
- Publikační typ
- časopisecké články 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 cholinesterázy * 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 cholinesterázy * MeSH
- sarin * MeSH
- VX MeSH Prohlížeč
In this review, the current progress in the research and development of butyrylcholinesterase (BChE) reactivators is summarised and the advantages or disadvantages of these reactivators are critically discussed. Organophosphorus compounds such as nerve agents (sarin, tabun, VX) or pesticides (chlorpyrifos, diazinon) cause irreversible inhibition of acetylcholinesterase (AChE) and BChE in the human body. While AChE inhibition can be life threatening due to cholinergic overstimulation and crisis, selective BChE inhibition has presumably no adverse effects. Because BChE is mostly found in plasma, its activity is important for the scavenging of organophosphates before they can reach AChE in the central nervous system. Therefore, this enzyme in combination with its reactivator can be used as a pseudo-catalytic scavenger of organophosphates. Three structural types of BChE reactivators were found, i.e. bisquaternary salts, monoquaternary salts and uncharged compounds. Although the reviewed reactivators have certain limitations, the promising candidates for BChE reactivation were found in each structural group.
- Klíčová slova
- Butyrylcholinesterase, Nerve agent, Oxime, Reactivator, Scavenger,
- MeSH
- acetylcholinesterasa metabolismus chemie MeSH
- butyrylcholinesterasa * metabolismus chemie MeSH
- cholinesterasové inhibitory * chemie farmakologie chemická syntéza MeSH
- lidé MeSH
- molekulární struktura MeSH
- organofosforové sloučeniny * chemie farmakologie MeSH
- reaktivátory cholinesterázy farmakologie chemie chemická syntéza MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Názvy látek
- acetylcholinesterasa MeSH
- butyrylcholinesterasa * MeSH
- cholinesterasové inhibitory * MeSH
- organofosforové sloučeniny * MeSH
- reaktivátory cholinesterázy MeSH
Enzyme handling and utilization bears many challenges such as their limited stability, intolerance of organic solvents, high cost, or inability to reuse. Most of these limitations can be overcome by enzyme immobilization on the surface of solid support. In this work, the recombinant form of human cholinesterases and monoamine oxidases as important drug targets for neurological diseases were immobilized on the surface of magnetic non-porous microparticles by a non-covalent bond utilizing the interaction between a His-tag terminus on the recombinant enzymes and cobalt (Co2+) ions immobilized on the magnetic microparticles. This type of binding led to targeted enzyme orientation, which completely preserved the catalytic activity and allowed high reproducibility of immobilization. In comparison with free enzymes, the immobilized enzymes showed exceptional stability in time and the possibility of repeated use. Relevant Km, Vmax, and IC50 values using known inhibitors were obtained using particular immobilized enzymes. Such immobilized enzymes on magnetic particles could serve as an excellent tool for a sustainable approach in the early stage of drug discovery.
- Klíčová slova
- Cholinesterases, Immobilization, Magnetic microparticles, Monoamine oxidases, Neurological disorders, Sustainability,
- MeSH
- acetylcholinesterasa metabolismus chemie MeSH
- analýza nákladů a výnosů MeSH
- enzymy imobilizované * chemie metabolismus MeSH
- kobalt * chemie MeSH
- lidé MeSH
- monoaminoxidasa metabolismus chemie MeSH
- nemoci nervového systému farmakoterapie enzymologie MeSH
- objevování léků * MeSH
- rekombinantní proteiny chemie metabolismus MeSH
- stabilita enzymů MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- acetylcholinesterasa MeSH
- enzymy imobilizované * MeSH
- kobalt * MeSH
- monoaminoxidasa MeSH
- rekombinantní proteiny MeSH
