RESUMEN
Injectable anticoagulants are widely used in medical procedures to prevent unwanted blood clotting. However, many lack safe, effective reversal agents. Here, we present new data on a previously described RNA origami-based, direct thrombin inhibitor (HEX01). We describe a new, fast-acting, specific, single-molecule reversal agent (antidote) and present in vivo data for the first time, including efficacy, reversibility, preliminary safety, and initial biodistribution studies. HEX01 contains multiple thrombin-binding aptamers appended on an RNA origami. It exhibits excellent anticoagulation activity in vitro and in vivo. The new single-molecule, DNA antidote (HEX02) reverses anticoagulation activity of HEX01 in human plasma within 30 s in vitro and functions effectively in a murine liver laceration model. Biodistribution studies of HEX01 in whole mice using ex vivo imaging show accumulation mainly in the liver over 24 h and with 10-fold lower concentrations in the kidneys. Additionally, we show that the HEX01/HEX02 system is non-cytotoxic to epithelial cell lines and non-hemolytic in vitro. Furthermore, we found no serum cytokine response to HEX01/HEX02 in a murine model. HEX01 and HEX02 represent a safe and effective coagulation control system with a fast-acting, specific reversal agent showing promise for potential drug development.
Asunto(s)
Aptámeros de Nucleótidos , Trombina , Animales , Ratones , Humanos , Aptámeros de Nucleótidos/farmacología , Aptámeros de Nucleótidos/química , Trombina/metabolismo , Coagulación Sanguínea/efectos de los fármacos , Distribución Tisular , ARN , Modelos Animales de Enfermedad , Hígado/metabolismo , Hígado/efectos de los fármacos , Anticoagulantes/farmacología , Anticoagulantes/química , Antitrombinas/farmacología , Antídotos/farmacología , Antídotos/químicaRESUMEN
Rising global population and increased food demands have resulted in the increased use of organophosphate pesticides (OPs), leading to toxin accumulation and transmission to humans. Pralidoxime (2-PAM), an FDA-approved drug, serves as an antidote for OP therapy. However, the atomic-level detoxification mechanisms regarding the design of novel antidotes remain unclear. This is the first study to examine the binding and unbinding pathways of 2-PAM to human acetylcholinesterase (HuAChE) through three identified doors using an enhanced sampling method called ligand-binding parallel cascade selection molecular dynamics (LB-PaCS-MD). Remarkably, LB-PaCS-MD could identify a predominant in-line binding mechanism through the acyl door at 63.79% ± 6.83%, also implicating it in a potential unbinding route (90.14% ± 4.22%). Interestingly, crucial conformational shifts in key residues, W86, Y341, and Y449, and the Ω loop significantly affect door dynamics and ligand binding modes. The LB-PaCS-MD technique can study ligand-binding pathways, thereby contributing to the design of antidotes and covalent drugs.
Asunto(s)
Acetilcolinesterasa , Inhibidores de la Colinesterasa , Simulación de Dinámica Molecular , Humanos , Acetilcolinesterasa/metabolismo , Acetilcolinesterasa/química , Antídotos/química , Antídotos/farmacología , Antídotos/metabolismo , Sitios de Unión , Inhibidores de la Colinesterasa/química , Inhibidores de la Colinesterasa/metabolismo , Inhibidores de la Colinesterasa/farmacología , Ligandos , Compuestos de Pralidoxima/química , Compuestos de Pralidoxima/metabolismo , Compuestos de Pralidoxima/farmacología , Unión ProteicaRESUMEN
Olesoxime, a cholesterol derivative with an oxime group, possesses the ability to cross the blood-brain barrier, and has demonstrated excellent safety and tolerability properties in clinical research. These characteristics indicate it may serve as a centrally active ligand of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), whose disruption of activity with organophosphate compounds (OP) leads to uncontrolled excitation and potentially life-threatening symptoms. To evaluate olesoxime as a binding ligand and reactivator of human AChE and BChE, we conducted in vitro kinetic studies with the active metabolite of insecticide parathion, paraoxon, and the warfare nerve agents sarin, cyclosarin, tabun, and VX. Our results showed that both enzymes possessed a binding affinity for olesoxime in the mid-micromolar range, higher than the antidotes in use (i.e., 2-PAM, HI-6, etc.). While olesoxime showed a weak ability to reactivate AChE, cyclosarin-inhibited BChE was reactivated with an overall reactivation rate constant comparable to that of standard oxime HI-6. Moreover, in combination with the oxime 2-PAM, the reactivation maximum increased by 10-30% for cyclosarin- and sarin-inhibited BChE. Molecular modeling revealed productive interactions between olesoxime and BChE, highlighting olesoxime as a potentially BChE-targeted therapy. Moreover, it might be added to OP poisoning treatment to increase the efficacy of BChE reactivation, and its cholesterol scaffold could provide a basis for the development of novel oxime antidotes.
Asunto(s)
Acetilcolinesterasa , Butirilcolinesterasa , Humanos , Butirilcolinesterasa/metabolismo , Butirilcolinesterasa/química , Acetilcolinesterasa/metabolismo , Acetilcolinesterasa/química , Ligandos , Oximas/química , Oximas/farmacología , Reactivadores de la Colinesterasa/farmacología , Reactivadores de la Colinesterasa/química , Inhibidores de la Colinesterasa/farmacología , Inhibidores de la Colinesterasa/química , Colestenonas/farmacología , Colestenonas/química , Cinética , Sarín/química , Proteínas Ligadas a GPI/metabolismo , Proteínas Ligadas a GPI/química , Proteínas Ligadas a GPI/antagonistas & inhibidores , Antídotos/farmacología , Antídotos/química , Colesterol/metabolismo , Colesterol/química , Compuestos OrganofosforadosRESUMEN
Thrombosis leads to elevated mortality rates and substantial medical expenses worldwide. Human factor IXa (HFIXa) protease is pivotal in tissue factor (TF)-mediated thrombin generation, and represents a promising target for anticoagulant therapy. We herein isolated novel DNA aptamers that specifically bind to HFIXa through systematic evolution of ligands by exponential enrichment (SELEX) method. We identified two distinct aptamers, seq 5 and seq 11, which demonstrated high binding affinity to HFIXa (Kd = 74.07 ± 2.53 nM, and 4.93 ± 0.15 nM, respectively). Computer software was used for conformational simulation and kinetic analysis of DNA aptamers and HFIXa binding. These aptamers dose-dependently prolonged activated partial thromboplastin time (aPTT) in plasma. We further rationally optimized the aptamers by truncation and site-directed mutation, and generated the truncated forms (Seq 5-1t, Seq 11-1t) and truncated-mutated forms (Seq 5-2tm, Seq 11-2tm). They also showed good anticoagulant effects. The rationally and structurally designed antidotes (seq 5-2b and seq 11-2b) were competitively bound to the DNA aptamers and effectively reversed the anticoagulant effect. This strategy provides DNA aptamer drug-antidote pair with effective anticoagulation and rapid reversal, developing advanced therapies by safe, regulatable aptamer drug-antidote pair.
Asunto(s)
Antídotos , Aptámeros de Nucleótidos , Factor IXa , Aptámeros de Nucleótidos/química , Aptámeros de Nucleótidos/farmacología , Humanos , Factor IXa/antagonistas & inhibidores , Factor IXa/metabolismo , Antídotos/farmacología , Antídotos/química , Antídotos/síntesis química , Relación Dosis-Respuesta a Droga , Anticoagulantes/farmacología , Anticoagulantes/química , Relación Estructura-Actividad , Estructura Molecular , Técnica SELEX de Producción de AptámerosRESUMEN
Thrombosis, the formation of blood clots within a blood vessel, can lead to severe complications including pulmonary embolism, cardiac arrest, and stroke. The most widely administered class of anticoagulants is heparin-based anticoagulants such as unfractionated heparin, low-molecular weight heparins (LMWHs), and fondaparinux. Protamine is the only FDA-approved heparin antidote. Protamine has limited efficacy neutralizing LMWHs and no reversal activity against fondaparinux. The use of protamine can lead to complications, including excessive bleeding, hypotension, and hypersensitivity, and has narrow therapeutic window. In this work, a new concept in the design of a universal heparin antidote: switchable protonation of cationic ligands, is presented. A library of macromolecular polyanion inhibitors (MPIs) is synthesized and screened to identify molecules that can neutralize all heparins with high selectivity and reduced toxicity. MPIs are developed by assembling cationic binding groups possessing switchable protonation states onto a polymer scaffold. By strategically selecting the identity and modulating the density of cationic binding groups on the polymer scaffold, a superior universal heparin reversal agent is developed with improved heparin-binding activity and increased hemocompatibility profiles leading to minimal effect on hemostasis. The activity of this heparin antidote is demonstrated using in vitro and in vivo studies.
Asunto(s)
Cationes , Heparina , Animales , Heparina/química , Heparina/farmacología , Ligandos , Cationes/química , Antagonistas de Heparina/química , Antagonistas de Heparina/farmacología , Humanos , Polielectrolitos/química , Polímeros/química , Antídotos/química , Antídotos/farmacología , Anticoagulantes/química , Anticoagulantes/farmacología , Ratones , Protaminas/química , Protaminas/farmacologíaRESUMEN
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.
Asunto(s)
Butirilcolinesterasa , Reactivadores de la Colinesterasa , Intoxicación por Organofosfatos , Oximas , Oximas/química , Oximas/farmacología , Reactivadores de la Colinesterasa/química , Reactivadores de la Colinesterasa/farmacología , Butirilcolinesterasa/metabolismo , Butirilcolinesterasa/química , Humanos , Intoxicación por Organofosfatos/tratamiento farmacológico , Acetilcolinesterasa/metabolismo , Antídotos/química , Antídotos/farmacología , Cinética , Inhibidores de la Colinesterasa/química , Inhibidores de la Colinesterasa/farmacología , Animales , Compuestos Organofosforados/químicaRESUMEN
The castor plant (Ricinus communis) is primarily known for its seeds, which contain a unique fatty acid called ricinoleic acid with several industrial and commercial applications. Castor seeds also contain ricin, a toxin considered a chemical and biological warfare agent. Despite years of investigation, there is still no effective antidote or vaccine available. However, some progress has been made, and the development of an effective treatment may be on the horizon. To provide an updated overview of this issue, we have conducted a comprehensive review of the literature on the current state of research in the fight against ricin. This review is based on the reported research and aims to address the challenges faced by researchers, as well as highlight the most successful cases achieved thus far. Our goal is to encourage the scientific community to continue their efforts in this critical search.
Asunto(s)
Antídotos , Ricina , Ricina/antagonistas & inhibidores , Ricina/química , Humanos , Antídotos/química , Antídotos/farmacología , Sustancias para la Guerra Química/química , AnimalesRESUMEN
Organophosphate (OP) toxicity is related to inhibition of acetylcholinesterase (AChE) activity, which plays a key role in the neurotransmission process. In this work, we report the ability of different zinc zeolitic imidazolate frameworks (ZIFs) to behave as potential antidotes against OP poisoning. The Zn-L coordination bond (L = purine, benzimidazole, imidazole, or 2-methylimidazole) is sensitive to the G-type nerve agent model compounds diisopropylfluorophosphate (DIFP) and diisopropylchlorophosphate, leading to P-X (X = F or Cl) bond breakdown into nontoxic diisopropylphosphate. P-X hydrolysis is accompanied by ZIF structural degradation (Zn-imidazolate bond hydrolysis), with the concomitant release of the imidazolate linkers and zinc ions representing up to 95% of ZIF particle dissolution. The delivered imidazolate nucleophilic attack on the OP@AChE adduct gives rise to the recovery of AChE enzymatic function. P-X bond breakdown, ZIF structural degradation, and AChE reactivation are dependent on imidazolate linker nucleophilicity, framework topology, and particle size. The best performance is obtained for 20 nm nanoparticles (NPs) of Zn(2-methylimidazolate)2 (sod ZIF-8) exhibiting a DIFP degradation half-life of 2.6 min and full recovery of AChE activity within 1 h. 20 nm sod ZIF-8 NPs are not neurotoxic, as proven by in vitro neuroblastoma cell culture viability tests.
Asunto(s)
Acetilcolinesterasa , Zeolitas , Acetilcolinesterasa/química , Organofosfatos/toxicidad , Zeolitas/química , Antídotos/química , Compuestos Orgánicos , Zinc/químicaRESUMEN
Owing to their potential to cause serious adverse health effects, significant efforts have been made to develop antidotes for organophosphate (OP) anticholinesterases, such as nerve agents. To be optimally effective, antidotes must not only reactivate inhibited target enzymes, but also have the ability to cross the blood-brain barrier (BBB). Progress has been made toward brain-penetrating acetylcholinesterase reactivators through the development of a new group of substituted phenoxyalkyl pyridinium oximes. To help in the selection and prioritization of compounds for future synthesis and testing within this class of chemicals, and to identify candidate broad-spectrum molecules, an in silico framework was developed to systematically generate structures and screen them for reactivation efficacy and BBB penetration potential.
Asunto(s)
Antídotos , Reactivadores de la Colinesterasa , Antídotos/farmacología , Antídotos/química , Inhibidores de la Colinesterasa/farmacología , Inhibidores de la Colinesterasa/química , Reactivadores de la Colinesterasa/farmacología , Reactivadores de la Colinesterasa/química , Organofosfatos , Acetilcolinesterasa/química , Oximas/químicaRESUMEN
The uncharged 3-hydroxy-2-pyridine aldoximes with protonatable tertiary amines are studied as antidotes in toxic organophosphates (OP) poisoning. Due to some of their specific structural features, we hypothesize that these compounds could exert diverse biological activity beyond their main scope of application. To examine this further, we performed an extensive cell-based assessment to determine their effects on human cells (SH-SY5Y, HEK293, HepG2, HK-2, myoblasts and myotubes) and possible mechanism of action. As our results indicated, aldoxime having a piperidine moiety did not induce significant toxicity up to 300 µM within 24 h, while those with a tetrahydroisoquinoline moiety, in the same concentration range, showed time-dependent effects and stimulated mitochondria-mediated activation of the intrinsic apoptosis pathway through ERK1/2 and p38-MAPK signaling and subsequent activation of initiator caspase 9 and executive caspase 3 accompanied with DNA damage as observed already after 4 h exposure. Mitochondria and fatty acid metabolism were also likely targets of 3-hydroxy-2-pyridine aldoximes with tetrahydroisoquinoline moiety, due to increased phosphorylation of acetyl-CoA carboxylase. In silico analysis predicted kinases as their most probable target class, while pharmacophores modeling additionally predicted the inhibition of a cytochrome P450cam. Overall, if the absence of significant toxicity for piperidine bearing aldoxime highlights the potential of its further studies in medical counter-measures, the observed biological activity of aldoximes with tetrahydroisoquinoline moiety could be indicative for future design of compounds either in a negative context in OP antidotes design, or in a positive one for design of compounds for the treatment of other phenomena like cell proliferating malignancies.
Asunto(s)
Neuroblastoma , Tetrahidroisoquinolinas , Humanos , Antídotos/química , Células HEK293 , Oximas/toxicidad , Oximas/química , Organofosfatos/química , Piridinas , Apoptosis , Transducción de Señal , Piperidinas , Tetrahidroisoquinolinas/toxicidadRESUMEN
Treatment for acute brain conditions remains a major challenge owing to the unavailability of antidotes, especially for organophosphorus compounds, exposure to which leads to rapid death. Despite recent advances in brain-targeted nano delivery systems (BTNDS), the traditional ones which have been developed will likely not lead to the quick release of an antidote, which is essential to counteract fast neurotoxic effects. Herein, we present a BTNDS using thermosensitive liposomes, without the need for functionalization, to obtain a platform for brain-targeted delivery, which has a simple structure and thus can be easily synthesized and scaled-up. The brain-targeting effect of BTNDS was amplified by phospholipase A2 (PLA2), an inflammatory biomarker. The combination of PLA2 and BTNDS significantly improved brain targeting, leading to an excellent emergency rescue effect - 83- and 4.8-fold better cerebral AChE reactivation response and survival time, respectively. These findings provide a promising strategy to generate a facile, druggable, and effective BTNDS.
Asunto(s)
Sistema de Administración de Fármacos con Nanopartículas , Oximas , Antídotos/química , Antídotos/farmacología , Encéfalo , Fosfolipasas A2RESUMEN
Organophosphorus compounds (OP) make up an important class of inhibitors, mostly employed as pesticides, even as chemical weapons. These toxic substances act through the inhibition of the acetylcholinesterase (AChE) enzyme, which results in elevated synaptic acetylcholine (ACh) levels, leading to serious adverse effects under the cholinergic syndrome. Many reactivators have been developed to combat the toxic effects of these AChE inhibitors. In this line, the oximes highlight because of their good reactivating power of cholinesterase enzymes. To date, no universal antidotes can reactivate AChE inhibited by any OP agent. This review summarizes the intoxication process by neurotoxic OP agents, along with the development of reactivators capable of reversing their effects, approaching aspects like the therapeutic and toxicological profile of these antidotes. Computational methods and conscious in vitro studies, capable of significantly predicting the toxicological profile of these drug candidates, might support the process of development of these reactivators before entering in vivo studies in animals, and then clinical trials. These approaches can assist in the design of safer and more effective molecules, reducing related cost and time for the process.
Asunto(s)
Antídotos , Reactivadores de la Colinesterasa , Animales , Antídotos/farmacología , Antídotos/uso terapéutico , Antídotos/química , Acetilcolinesterasa/química , Reactivadores de la Colinesterasa/uso terapéutico , Reactivadores de la Colinesterasa/toxicidad , Compuestos Organofosforados , Oximas/uso terapéutico , Oximas/toxicidad , Inhibidores de la Colinesterasa/toxicidadRESUMEN
Organophosphorus compounds (OPs) pose great military and civilian hazards. However, therapeutic and prophylactic antidotes against OP poisoning remain challenging. In this study, we first developed a novel nanoscavenger (rOPH/ZIF-8@E-Lipo) against methyl paraoxon (MP) poisoning using enzyme immobilization and erythrocyte-liposome hybrid membrane camouflage techniques. Then, we evaluated the physicochemical characterization, stability, and biocompatibility of the nanoscavengers. Afterward, we examined acetylcholinesterase (AChE) activity, cell viability, and intracellular reactive oxygen species (ROS) to indicate the protective effects of the nanoscavengers in vitro. Following the pharmacokinetic and biodistribution studies, we further evaluated the therapeutic and prophylactic detoxification efficacy of the nanoscavengers against MP in various poisoning settings. Finally, we explored the penetration capacity of the nanoscavengers across the blood-brain barrier (BBB). The present study validated the successful construction of a novel nanoscavenger with excellent stability and biocompatibility. In vitro, the resulting nanoscavenger exhibited a significant protection against MP-induced AChE inactivation, oxidative stress, and cytotoxicity. In vivo, apart from the positive therapeutic effects, the nanoscavengers also exerted significant prophylactic detoxification efficacy against single lethal MP exposure, repeated lethal MP challenges, and sublethal MP poisoning. These excellent detoxification effects of the nanoscavengers against OPs may originate from a dual-mode mechanism of inner recombinant organophosphorus hydrolase (rOPH) and outer erythrocyte membrane-anchored AChE. Finally, in vitro and in vivo studies jointly demonstrated that monosialoganglioside (GM1)-modified rOPH/ZIF-8@E-Lipo could penetrate the BBB with high efficiency. In conclusion, a stable and safe dual-modal nanoscavenger was developed with BBB penetration capability, providing a promising strategy for the treatment and prevention of OP poisoning.
Asunto(s)
Acetilcolinesterasa , Compuestos Organofosforados , Acetilcolinesterasa/metabolismo , Antídotos/química , Antídotos/farmacología , Antídotos/uso terapéutico , Arildialquilfosfatasa , Inhibidores de la Colinesterasa/farmacología , Gangliósido G(M1) , Liposomas , Paraoxon/análogos & derivados , Especies Reactivas de Oxígeno , Distribución TisularRESUMEN
The effectiveness of an amphoteric cryogel (AAC) as an oral sorbent (enerosorbent) for the treatment of acute poisoning of small animals (rats) with heavy metals (HMs) was studied in in vivo experiments. The morphological structure of the cryogel was examined using scanning electron microscopy/energy-dispersive X-ray analysis and confocal microscopy. The use of the cryogel in the treatment of rats administered an LD50 dose of Cd(NO3)2, CsNO3, Sr(NO3)2, or HgCl2 in aqueous solution showed their high survival rate compared to the control group, which did not receive such treatment. The histological and chemical analysis of internal tissues and the biochemical analysis of the blood of the experimental animals showed the effectiveness of the cryogel in protecting the animals against the damaging effect of HMs on the organism comparable with unithiol, a chelating agent based on 2,3-dimercapto-1-propane sulfonic acid sodium salt (DMPS) approved for the treatment of acute poisoning with some heavy metals.
Asunto(s)
Antídotos , Quelantes , Criogeles , Intoxicación por Metales Pesados/tratamiento farmacológico , Animales , Antídotos/síntesis química , Antídotos/química , Antídotos/farmacología , Quelantes/síntesis química , Quelantes/química , Quelantes/farmacología , Criogeles/síntesis química , Criogeles/química , Criogeles/farmacología , Intoxicación por Metales Pesados/metabolismo , Masculino , Metales Pesados/metabolismo , RatasRESUMEN
Although great potential hazards and threats still occur from sulfur mustard, there are no specific medicine or therapy for the intoxication of sulfur mustard. Herein, we have demonstrated a supramolecular approach for the detoxification of the sulfur mustard simulant CEES (4) in vitro and in vivo by carboxylatopillar[5]arene potassium salts (CP[5]AK 1) efficiently based on host-guest interactions. The encapsulation of CEES (4) by the cavity of the pillar[5]arene 2 is driven by C-H···π interactions between CEES (4) and the electron-rich cavity of pillar[5]arene 2, which was investigated by 1H NMR titration, density functional theory studies, and the independent gradient model studies. CEES (4) is degradated to the reactive sulfonium salts quickly in aqueous media, resulting in the alkylation of DNA and proteins. The sulfonium salts can be encapsulated by CP[5]AK 1 efficiently, which accelerates the degradation of the sulfonium salts about 14 times. The cell and animal experiments indicated that the bioactivities of the sulfonium salts are inhibited with the formation of stable host-guest complexes, and CP[5]AK 1 has a good therapeutic effect on the damages caused by CEES (4) at either pre- or post-treatments. Due to the low cytotoxicity and good therapeutic effect, the anionic pillar[5]arenes are expected to be developed as specific antidotes against sulfur mustard (HD).
Asunto(s)
Antídotos , Gas Mostaza , Animales , Humanos , Ratas , Antídotos/química , Antídotos/farmacología , Supervivencia Celular/efectos de los fármacos , Teoría Funcional de la Densidad , Oftalmopatías/tratamiento farmacológico , Oftalmopatías/patología , Células HEK293 , Sustancias Macromoleculares/química , Sustancias Macromoleculares/farmacología , Ensayo de Materiales , Estructura Molecular , Gas Mostaza/química , Gas Mostaza/metabolismo , Gas Mostaza/toxicidad , Enfermedades Respiratorias/tratamiento farmacológico , Enfermedades Respiratorias/patología , Sales (Química)/química , Sales (Química)/metabolismo , Sales (Química)/toxicidadRESUMEN
Poisoning is a leading cause of admission to medical emergency departments and intensive care units. Supramolecular detoxification, which involves injecting supramolecular receptors that bind with toxins to suppress their biological activity, is an emerging strategy for poisoning treatment; it has few requirements and a broad application scope. However, it is still a formidable challenge to design supramolecular therapeutic materials as an antidote to macromolecular toxins, because the large size, flexible conformation, and presence of multiple and diverse binding sites of biomacromolecules hinder their recognition. Herein, a supramolecular antidote to macromolecular toxins is developed through the coassembly of macrocyclic amphiphiles, relying on heteromultivalent recognition between the coassembled components and toxic macromolecules. The coassembly of amphiphilic cyclodextrin and calixarene strongly and selectively captures melittin, a toxin studied herein; this imparts various therapeutic effects such as inhibiting the interactions of melittin with cell membranes, alleviating melittin cytotoxicity and hemolytic toxicity, reducing the mortality rate of melittin-poisoned mice, and mitigating damage to major organs. The use of the proposed antidote overcomes the limitation of supramolecular detoxification applicability to only small-molecular toxins. The antidote can also detoxify other macromolecular toxins as long as selective and strong binding is achieved because of the coassembling tunability.
Asunto(s)
Antídotos/química , Sustancias Macromoleculares/química , Meliteno/química , Animales , Antídotos/metabolismo , Antídotos/farmacología , Péptidos Catiónicos Antimicrobianos/química , Péptidos Catiónicos Antimicrobianos/farmacología , Membrana Celular/química , Supervivencia Celular/efectos de los fármacos , Ciclodextrinas/química , Células HEK293 , Hemólisis/efectos de los fármacos , Humanos , Hígado/efectos de los fármacos , Hígado/patología , Sustancias Macromoleculares/metabolismo , Meliteno/metabolismo , Meliteno/farmacología , Ratones , Venenos de Araña/química , Venenos de Araña/farmacologíaRESUMEN
Gelsenicine is one of the most toxic compounds in the genus Gelsemium, but the mechanism of toxicity is not clear. In this paper, tandem mass tag quantitative phosphoproteomics was used to study the changes in protein phosphorylation in different brain regions at different time points after gelsenicine poisoning in mice. The correlation between neurotransmitter receptors and the toxicity of gelsenicine was analyzed by molecular docking and rescue experiments. Parallel reaction monitoring (PRM) was used to verify the related proteins. A total of 17877 unique phosphosites were quantified and mapped to 4170 brain proteins to understand the signaling pathways. Phosphoproteomics revealed gelsenicine poisoning mainly affected protein phosphorylation levels in the hippocampus, and through bioinformatics analysis, it was found gelsenicine poisoning significantly affected neurotransmitter synaptic pathway. The molecular docking results showed that gelsenicine could bind to the N-methyl-D-aspartic acid receptor (NMDAR). In addition, we found that NMDA was effective in improving the survival rate of the animals tested, and this effect was associated with reduced protein phosphorylation by PRM validation. The results revealed that gelsenicine affects neurotransmitter release and receptor function. This is the first demonstration that NMDA receptor-mediated excitotoxicity is a key signaling pathway in the toxicity of gelsenicine.
Asunto(s)
Alcaloides Indólicos/toxicidad , Proteómica/métodos , Receptores de N-Metil-D-Aspartato/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , Antídotos/química , Antídotos/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos ICR , Modelos Moleculares , Simulación del Acoplamiento Molecular , N-Metilaspartato/farmacología , Conformación Proteica , Receptores de N-Metil-D-Aspartato/agonistas , Receptores de N-Metil-D-Aspartato/genéticaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Theriac is considered the most popular cure-all multi-ingredient medicine and has been used for more than two millennia. It has also been used as one of the most important anti-epidemic drugs up to the 19th c., treated as an emergency medicine in case of e.g. bubonic plague. AIM OF THE STUDY: Until now, no reliable information regarding the pharmacological effect of the treacle was available, including its possible toxic or narcotic properties. In order to change the state of knowledge in this matter we have selected the Theriac recipe that had been actually used for producing the treacle in 1630, which was confirmed by the official municipal documents of the time. METHODS: The recipe was written in Latin, with the use of pre-Linnean nomenclature and then apothecary common names, which required translation into the modern scientific language in order to get reliable pharmacological conclusions. The information from historical sources has been compiled with the pharmacological data concerning the most potent compounds, which for the first time made it possible to calculate the amounts of active compounds in the doses taken by then patients. RESULTS: Only two species included in Theriac can be harmful in humans: poppy and sea squill, but in both cases the calculated quantity of morphine and cardiac glycosides, respectively, were below toxic level. There are no indications, both from the historical and pharmacological point of view, for Theriac being toxic or narcotic in patients, when used as prescribed. CONCLUSIONS: As for now, the most probable is that the treacle owed its postulated efficacy in the main indications to the placebo effect. Still, the results should be further confirmed by reconstructing the actual Theriac and subjecting it to modern tests and analyses.
Asunto(s)
Antídotos/historia , Antídotos/farmacología , Venenos , Charlatanería , Antídotos/química , Combinación de Medicamentos , Europa (Continente) , Historia del Siglo XV , Historia del Siglo XVI , Historia del Siglo XVII , Historia del Siglo XVIII , Historia del Siglo XIX , Historia Antigua , Historia Medieval , Humanos , Materia MedicaRESUMEN
Oximes, investigated as antidotes against organophosphates (OP) poisoning, are known to display toxic effects on a cellular level, which could be explained beyond action on acetylcholinesterase as their main target. To investigate this further, we performed an in vitro cell-based evaluation of effects of two structurally diverse oxime groups at concentrations of up to 800 µM, on several cell models: skeletal muscle, kidney, liver, and neural cells. As indicated by our results, compounds with an imidazolium core induced necrosis, unregulated cell death characterized by a cell burst, increased formation of reactive oxygen species, and activation of antioxidant scavenging. On the other hand, oximes with a pyridinium core activated apoptosis through specific caspases 3, 8, and/or 9. Interestingly, some of the compounds exhibited a synergistic effect. Moreover, we generated a pharmacophore model for each oxime series and identified ligands from public databases that map to generated pharmacophores. Several interesting hits were obtained including chemotherapeutics and specific inhibitors. We were able to define the possible structural features of tested oximes triggering toxic effects: chlorine atoms in combination with but-2(E)-en-1,4-diyl linker and adding a second benzene ring with substituents such as chlorine and/or methyl on the imidazolium core. Such oximes could not be used in further OP antidote development research, but could be introduced in other research studies on new specific targets. This could undoubtedly result in an overall improved wider use of unexplored oxime database created so far in OP antidotes field of research in a completely new perspective.
Asunto(s)
Antídotos/toxicidad , Oximas/toxicidad , Compuestos de Piridinio/toxicidad , Muerte Celular Regulada/efectos de los fármacos , Animales , Antídotos/química , Antioxidantes/metabolismo , Apoptosis/efectos de los fármacos , Línea Celular , Línea Celular Tumoral , Perros , Sinergismo Farmacológico , Humanos , Células de Riñón Canino Madin Darby , Oximas/administración & dosificación , Oximas/química , Compuestos de Piridinio/química , Especies Reactivas de Oxígeno/metabolismo , Relación Estructura-ActividadRESUMEN
Protein-protein (e.g., antibody-antigen) interactions comprise multiple weak interactions. We have previously reported that lipid nanoparticles (LNPs) bind to and neutralize target toxic peptides after multifunctionalization of the LNP surface (MF-LNPs) with amino acid derivatives that induce weak interactions; however, the MF-LNPs aggregated after target capture and showed short blood circulation times. Here we optimized polyethylene glycol (PEG)-modified MF-LNPs (PEG-MF-LNPs) to inhibit the aggregation and increase the blood circulation time. Melittin was used as a target toxin, and MF-LNPs were prepared with negatively charged, hydrophobic, and neutral amino-acid-derivative-conjugated functional lipids. In this study, MF-LNPs modified with only PEG5k (PEG5k-MF-LNPs) and with both PEG5k and PEG2k (PEGmix-MF-LNPs) were prepared, where PEG5k and PEG2k represent PEG with a molecular weight of 5000 and 2000, respectively. PEGylation of the MF-LNPs did not decrease the melittin neutralization ability of nonPEGylated MF-LNPs, as tested by hemolysis assay. The PEGmix-MF-LNPs showed better blood circulation characteristics than the PEG5k-MF-LNPs. Although the nonPEGylated MF-LNPs immediately aggregated when mixed with melittin, the PEGmix-MF-LNPs did not aggregate. The PEGmix-MF-LNPs dramatically increased the survival rate of melittin-treated mice, whereas the nonPEGylated MF-LNPs increased slightly. These results provide a fundamental strategy to improve the in vivo toxin neutralization ability of MF-LNPs.