RESUMO
The Warburg effect is a hallmark of cancer that refers to the preference of cancer cells to metabolize glucose anaerobically rather than aerobically1,2. This results in substantial accumulation of lacate, the end product of anaerobic glycolysis, in cancer cells3. However, how cancer metabolism affects chemotherapy response and DNA repair in general remains incompletely understood. Here we report that lactate-driven lactylation of NBS1 promotes homologous recombination (HR)-mediated DNA repair. Lactylation of NBS1 at lysine 388 (K388) is essential for MRE11-RAD50-NBS1 (MRN) complex formation and the accumulation of HR repair proteins at the sites of DNA double-strand breaks. Furthermore, we identify TIP60 as the NBS1 lysine lactyltransferase and the 'writer' of NBS1 K388 lactylation, and HDAC3 as the NBS1 de-lactylase. High levels of NBS1 K388 lactylation predict poor patient outcome of neoadjuvant chemotherapy, and lactate reduction using either genetic depletion of lactate dehydrogenase A (LDHA) or stiripentol, a lactate dehydrogenase A inhibitor used clinically for anti-epileptic treatment, inhibited NBS1 K388 lactylation, decreased DNA repair efficacy and overcame resistance to chemotherapy. In summary, our work identifies NBS1 lactylation as a critical mechanism for genome stability that contributes to chemotherapy resistance and identifies inhibition of lactate production as a promising therapeutic cancer strategy.
Assuntos
Proteínas de Ciclo Celular , Resistencia a Medicamentos Antineoplásicos , Ácido Láctico , Proteínas Nucleares , Reparo de DNA por Recombinação , Animais , Feminino , Humanos , Masculino , Camundongos , Hidrolases Anidrido Ácido/metabolismo , Anaerobiose , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Quebras de DNA de Cadeia Dupla , Proteínas de Ligação a DNA/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Instabilidade Genômica , Ácido Láctico/metabolismo , Lisina/química , Lisina/metabolismo , Lisina Acetiltransferase 5/metabolismo , Lisina Acetiltransferase 5/genética , Proteína Homóloga a MRE11/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/genética , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Organoides , Glicólise , Terapia Neoadjuvante , L-Lactato Desidrogenase/antagonistas & inibidores , L-Lactato Desidrogenase/deficiência , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Anticonvulsivantes/farmacologiaRESUMO
Epilepsy remains a major health concern as anti-seizure medications frequently fail, and there is currently no treatment to stop or prevent epileptogenesis, the process underlying the onset and progression of epilepsy. The identification of the pathological processes underlying epileptogenesis is instrumental to the development of drugs that may prevent the generation of seizures or control pharmaco-resistant seizures, which affect about 30% of patients. mTOR signalling and neuroinflammation have been recognized as critical pathways that are activated in brain cells in epilepsy. They represent a potential node of biological convergence in structural epilepsies with either a genetic or an acquired aetiology. Interventional studies in animal models and clinical studies give strong support to the involvement of each pathway in epilepsy. In this Review, we focus on available knowledge about the pathophysiological features of mTOR signalling and the neuroinflammatory brain response, and their interactions, in epilepsy. We discuss mitigation strategies for each pathway that display therapeutic effects in experimental and clinical epilepsy. A deeper understanding of these interconnected molecular cascades could enhance our strategies for managing epilepsy. This could pave the way for new treatments to fill the gaps in the development of preventative or disease-modifying drugs, thus overcoming the limitations of current symptomatic medications.
Assuntos
Progressão da Doença , Epilepsia , Doenças Neuroinflamatórias , Serina-Treonina Quinases TOR , Humanos , Epilepsia/tratamento farmacológico , Animais , Serina-Treonina Quinases TOR/metabolismo , Doenças Neuroinflamatórias/tratamento farmacológico , Doenças Neuroinflamatórias/metabolismo , Transdução de Sinais/fisiologia , Encéfalo/metabolismo , Encéfalo/patologia , Anticonvulsivantes/uso terapêutico , Anticonvulsivantes/farmacologiaRESUMO
γ-Aminobutyric acid (GABA) transporter 1 (GAT1)1 regulates neuronal excitation of the central nervous system by clearing the synaptic cleft of the inhibitory neurotransmitter GABA upon its release from synaptic vesicles. Elevating the levels of GABA in the synaptic cleft, by inhibiting GABA reuptake transporters, is an established strategy to treat neurological disorders, such as epilepsy2. Here we determined the cryo-electron microscopy structure of full-length, wild-type human GAT1 in complex with its clinically used inhibitor tiagabine3, with an ordered part of only 60 kDa. Our structure reveals that tiagabine locks GAT1 in the inward-open conformation, by blocking the intracellular gate of the GABA release pathway, and thus suppresses neurotransmitter uptake. Our results provide insights into the mixed-type inhibition of GAT1 by tiagabine, which is an important anticonvulsant medication. Its pharmacodynamic profile, confirmed by our experimental data, suggests initial binding of tiagabine to the substrate-binding site in the outward-open conformation, whereas our structure presents the drug stalling the transporter in the inward-open conformation, consistent with a two-step mechanism of inhibition4. The presented structure of GAT1 gives crucial insights into the biology and pharmacology of this important neurotransmitter transporter and provides blueprints for the rational design of neuromodulators, as well as moving the boundaries of what is considered possible in single-particle cryo-electron microscopy of challenging membrane proteins.
Assuntos
Proteínas da Membrana Plasmática de Transporte de GABA , Inibidores da Captação de GABA , Ácido gama-Aminobutírico , Anticonvulsivantes/química , Anticonvulsivantes/farmacologia , Microscopia Crioeletrônica , Proteínas da Membrana Plasmática de Transporte de GABA/química , Proteínas da Membrana Plasmática de Transporte de GABA/metabolismo , Proteínas da Membrana Plasmática de Transporte de GABA/ultraestrutura , Inibidores da Captação de GABA/química , Inibidores da Captação de GABA/farmacologia , Humanos , Neurotransmissores/metabolismo , Conformação Proteica/efeitos dos fármacos , Tiagabina/química , Tiagabina/metabolismo , Tiagabina/farmacologia , Ácido gama-Aminobutírico/metabolismoRESUMO
Seizures and other forms of neurovolatility are emerging as druggable prodromal mechanisms that link traumatic brain injury (TBI) to the progression of later dementias. TBI neurotrauma has both acute and long-term impacts on health, and TBI is a leading risk factor for dementias, including chronic traumatic encephalopathy and Alzheimer's disease. Treatment of TBI already considers acute management of posttraumatic seizures and epilepsy, and impressive efforts have optimized regimens of antiepileptic drugs (AEDs) toward that goal. Here we consider that expanding these management strategies could determine which AED regimens best prevent dementia progression in TBI patients. Challenges with this prophylactic strategy include the potential consequences of prolonged AED treatment and that a large subset of patients are refractory to available AEDs. Addressing these challenges is warranted because the management of seizure activity following TBI offers a rare opportunity to prevent the onset or progression of devastating dementias.
Assuntos
Lesões Encefálicas Traumáticas , Demência , Epilepsia Pós-Traumática , Humanos , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Epilepsia Pós-Traumática/complicações , Epilepsia Pós-Traumática/tratamento farmacológico , Epilepsia Pós-Traumática/prevenção & controle , Lesões Encefálicas Traumáticas/tratamento farmacológico , Lesões Encefálicas Traumáticas/complicações , Convulsões/tratamento farmacológico , Convulsões/etiologia , Demência/tratamento farmacológico , Demência/prevenção & controleRESUMO
Voltage-gated sodium (Nav) channels govern membrane excitability, thus setting the foundation for various physiological and neuronal processes. Nav channels serve as the primary targets for several classes of widely used and investigational drugs, including local anesthetics, antiepileptic drugs, antiarrhythmics, and analgesics. In this study, we present cryogenic electron microscopy (cryo-EM) structures of human Nav1.7 bound to two clinical drugs, riluzole (RLZ) and lamotrigine (LTG), at resolutions of 2.9 Å and 2.7 Å, respectively. A 3D EM reconstruction of ligand-free Nav1.7 was also obtained at 2.1 Å resolution. RLZ resides in the central cavity of the pore domain and is coordinated by residues from repeats III and IV. Whereas one LTG molecule also binds to the central cavity, the other is found beneath the intracellular gate, known as site BIG. Therefore, LTG, similar to lacosamide and cannabidiol, blocks Nav channels via a dual-pocket mechanism. These structures, complemented with docking and mutational analyses, also explain the structure-activity relationships of the LTG-related linear 6,6 series that have been developed for improved efficacy and subtype specificity on different Nav channels. Our findings reveal the molecular basis for these drugs' mechanism of action and will aid the development of novel antiepileptic and pain-relieving drugs.
Assuntos
Canabidiol , Canais de Sódio Disparados por Voltagem , Humanos , Anticonvulsivantes/farmacologia , Lamotrigina/farmacologia , Sódio/metabolismo , Canais de Sódio Disparados por Voltagem/químicaRESUMO
Glial cells play relevant roles in neuroinflammation caused by epilepsy. Elevated hemichannel (HC) activity formed by connexins (Cxs) or pannexin1 (Panx1) largely explains brain dysfunctions commonly caused by neuroinflammation. Glia express HCs formed by Cxs 43, 30, or 26, while glia and neurons both express HCs formed by Panx1. Cx43 HCs allow for the influx of Ca2+, which promotes glial reactivity, enabling the release of the gliotransmitters that contribute to neuronal over-stimulation. Valproate (VPA), an antiseizure medication, has pleiotropic actions on neuronal molecular targets, and their action on glial cell HCs remains elusive. We used HeLa cells transfected with Cx43, Cx30, Cx26, or Panx1 to determine the effect of VPA on HC activity in the brain. VPA slightly increased HC activity under basal conditions, but significantly enhanced it in cells pre-exposed to conditions that promoted HC activity. Furthermore, VPA increased ATP release through Cx43 HCs. The increased HC activity caused by VPA was resistant to washout, being consistent with in silico studies, which predicted the binding site for VPA and Cx43, as well as for Panx1 HCs on the intracellular side, suggesting that VPA first enters through HCs, after which their activity increases.
Assuntos
Anticonvulsivantes , Conexinas , Ácido Valproico , Ácido Valproico/farmacologia , Humanos , Anticonvulsivantes/farmacologia , Conexinas/metabolismo , Células HeLa , Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Conexina 43/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Trifosfato de Adenosina/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Animais , Epilepsia/metabolismo , Epilepsia/tratamento farmacológico , Epilepsia/induzido quimicamenteRESUMO
There are limited therapeutic options for patients with Dravet syndrome (DS). The equilibrative nucleoside transporters 1 (ENT1) mediate both the influx and efflux of adenosine across the cell membrane exerted beneficial effects in the treatment of epilepsy. This study aimed to evaluate the anticonvulsant effect of the ENT1 inhibitor in an animal model of DS (Scn1aE1099X/+ mice). J7 (5 mg/kg) treatment was efficacious in elevating seizure threshold in Scn1aE1099X/+ mice after hyperthermia exposure. Moreover, the J7 treatment significantly reduced the frequency of spontaneous excitatory post-synaptic currents (sEPSCs, ~35% reduction) without affecting the amplitude in dentate gyrus (DG) granule cells. Pretreatment with the adenosine A1 receptor (A1R) antagonist, DPCPX, abolished the J7 effects on sEPSCs. These observations suggest that the J7 shows an anticonvulsant effect in hyperthermia-induced seizures in Scn1aE1099X/+ mice. This effect possibly acts on presynaptic A1R-mediated signaling modulation in granule cells.
Assuntos
Epilepsias Mioclônicas , Epilepsia , Humanos , Camundongos , Animais , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Nucleosídeos/uso terapêutico , Epilepsias Mioclônicas/tratamento farmacológico , Epilepsias Mioclônicas/genética , Epilepsias Mioclônicas/metabolismo , Neurônios/metabolismo , Modelos Animais de Doenças , Canal de Sódio Disparado por Voltagem NAV1.1/genéticaRESUMO
The R-type voltage-gated calcium channel CaV2.3 is predominantly located in the presynapse and is implicated in distinct types of epileptic seizures. It has consequently emerged as a molecular target in seizure treatment. Here, we determined the cryo-EM structure of the CaV2.3-α2δ1-ß1 complex in the topiramate-bound state at a 3.0 Å resolution. We provide a snapshot of the binding site of topiramate, a widely prescribed antiepileptic drug, on a voltage-gated ion channel. The binding site is located at an intracellular juxtamembrane hydrophilic cavity. Further structural analysis revealed that topiramate may allosterically facilitate channel inactivation. These findings provide fundamental insights into the mechanism underlying the inhibitory effect of topiramate on CaV and NaV channels, elucidating a previously unseen modulator binding site and thus pointing toward a route for the development of new drugs.
Assuntos
Anticonvulsivantes , Canais de Cálcio Tipo R , Microscopia Crioeletrônica , Topiramato , Anticonvulsivantes/química , Anticonvulsivantes/farmacologia , Topiramato/química , Topiramato/farmacologia , Humanos , Regulação Alostérica/efeitos dos fármacos , Canais de Cálcio Tipo R/química , Canais de Cálcio Tipo R/metabolismo , Sítios de Ligação , Modelos Moleculares , Células HEK293 , Conformação Proteica , Frutose/química , Frutose/análogos & derivados , Frutose/metabolismo , Animais , Proteínas de Transporte de CátionsRESUMO
Benzodiazepine pharmacoresistance develops when treatment of status epilepticus (SE) is delayed. This response may result from gamma-aminobutyric acid A receptors (GABAAR) internalization that follows prolonged SE; this receptor trafficking results in fewer GABAAR in the synapse to restore inhibition. Increase in synaptic N-methyl-D-aspartate receptors (NMDAR) also occurs in rodent models of SE. Lacosamide, a third-generation antiseizure medication (ASM), acts on the slow inactivation of voltage-gated sodium channels. Another ASM, rufinamide, similarly acts on sodium channels by extending the duration of time spent in the inactivation stage. Combination therapy of the benzodiazepine midazolam, NMDAR antagonist ketamine, and ASMs lacosamide (or rufinamide) was investigated for efficacy against soman (GD)-induced SE and neuropathology. Adult male rats implanted with telemetry transmitters for monitoring electroencephalographic (EEG) activity were exposed to a seizure-inducing dose of GD and treated with an admix of atropine sulfate and HI-6 1 minute later and with midazolam monotherapy or combination therapy 40 minutes after EEG seizure onset. Rats were monitored continuously for seizure activity for two weeks, after which brains were processed for assessment of neurodegeneration, neuronal loss, and neuroinflammatory responses. Simultaneous administration of midazolam, ketamine, and lacosamide (or rufinamide) was more protective against GD-induced SE compared with midazolam monotherapy. In general, lacosamide triple therapy had more positive outcomes on measures of epileptogenesis, EEG power integral, and the number of brain regions protected from neuropathology compared with rats treated with rufinamide triple therapy. Overall, both drugs were well tolerated in these combination models. SIGNIFICANCE STATEMENT: We currently report on improved efficacy of antiseizure medications lacosamide and rufinamide, each administered in combination with ketamine (NMDAR antagonist) and midazolam (benzodiazepine), in combatting soman (GD)-induced seizure, epileptogenesis, and brain pathology over that provided by midazolam monotherapy, or dual therapy of midazolam and lacosamide (or rufinamide) in rats. Administration of lacosamide as adjunct to midazolam and ketamine was particularly effective against GD-induced toxicity. However, protection was incomplete, suggesting the need for further study.
Assuntos
Ketamina , Soman , Estado Epiléptico , Triazóis , Ratos , Masculino , Animais , Midazolam/uso terapêutico , Midazolam/farmacologia , Lacosamida/efeitos adversos , Ketamina/farmacologia , Ketamina/uso terapêutico , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/tratamento farmacológico , Convulsões/tratamento farmacológico , Benzodiazepinas , Colinérgicos/efeitos adversos , Ácido gama-AminobutíricoRESUMO
Status epilepticus (SE) is a life-threatening development of self-sustaining seizures that becomes resistant to benzodiazepines when treatment is delayed. Benzodiazepine pharmacoresistance is thought in part to result from internalization of synaptic GABAA receptors, which are the main target of the drug. The naturally occurring neurosteroid allopregnanolone is a therapy of interest against SE for its ability to modulate all isoforms of GABAA receptors. Ketamine, an N-methyl-D-aspartate (NMDA) receptor antagonist, has been partially effective in combination with benzodiazepines in mitigating SE-associated neurotoxicity. In this study, allopregnanolone as an adjunct to midazolam or midazolam-ketamine combination therapy was evaluated for efficacy against cholinergic-induced SE. Adult male rats implanted with electroencephalographic (EEG) telemetry devices were exposed to the organophosphorus chemical (OP) soman (GD) and treated with an admix of atropine sulfate and HI-6 at 1 minute after exposure followed by midazolam, midazolam-allopregnanolone, or midazolam-ketamine-allopregnanolone 40 minutes after seizure onset. Neurodegeneration, neuronal loss, and neuroinflammation were assessed 2 weeks after GD exposure. Seizure activity, EEG power integral, and epileptogenesis were also compared among groups. Overall, midazolam-ketamine-allopregnanolone combination therapy was effective in reducing cholinergic-induced toxic signs and neuropathology, particularly in the thalamus and hippocampus. Higher dosage of allopregnanolone administered in combination with midazolam and ketamine was also effective in reducing EEG power integral and epileptogenesis. The current study reports that there is a promising potential of neurosteroids in combination with benzodiazepine and ketamine treatments in a GD model of SE. SIGNIFICANCE STATEMENT: Allopregnanolone, a naturally occurring neurosteroid, reduced pathologies associated with soman (GD) exposure such as epileptogenesis, neurodegeneration, and neuroinflammation, and suppressed GD-induced toxic signs when used as an adjunct to midazolam and ketamine in a delayed treatment model of soman-induced status epilepticus (SE) in rats. However, protection was incomplete, suggesting that further studies are needed to identify optimal combinations of antiseizure medications and routes of administration for maximal efficacy against cholinergic-induced SE.
Assuntos
Ketamina , Neuroesteroides , Soman , Estado Epiléptico , Ratos , Masculino , Animais , Midazolam/farmacologia , Midazolam/uso terapêutico , Ketamina/farmacologia , Ketamina/uso terapêutico , Pregnanolona/efeitos adversos , Soman/toxicidade , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Doenças Neuroinflamatórias , Neuroesteroides/uso terapêutico , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/tratamento farmacológico , Convulsões/tratamento farmacológico , Benzodiazepinas , Colinérgicos/efeitos adversos , Receptores de GABA-A , Ácido gama-AminobutíricoRESUMO
The development of refractory status epilepticus (SE) following sarin intoxication presents a therapeutic challenge. Here, we evaluated the efficacy of delayed combined double or triple treatment in reducing abnormal epileptiform seizure activity (ESA) and the ensuing long-term neuronal insult. SE was induced in rats by exposure to 1.2 LD50 sarin followed by treatment with atropine and TMB4 (TA) 1 min later. Double treatment with ketamine and midazolam or triple treatment with ketamine, midazolam and levetiracetam was administered 30 min post-exposure, and the results were compared to those of single treatment with midazolam alone or triple treatment with ketamine, midazolam, and valproate, which was previously shown to ameliorate this neurological insult. Toxicity and electrocorticogram activity were monitored during the first week, and behavioral evaluations were performed 2 weeks post-exposure, followed by biochemical and immunohistopathological analyses. Both double and triple treatment reduced mortality and enhanced weight recovery compared to TA-only treatment. Triple treatment and, to a lesser extent, double treatment significantly ameliorated the ESA duration. Compared to the TA-only or the TA+ midazolam treatment, both double and triple treatment reduced the sarin-induced increase in the neuroinflammatory marker PGE2 and the brain damage marker TSPO and decreased gliosis, astrocytosis and neuronal damage. Finally, both double and triple treatment prevented a change in behavior, as measured in the open field test. No significant difference was observed between the efficacies of the two triple treatments, and both triple combinations completely prevented brain injury (no differences from the naïve rats). Delayed double and, to a greater extent, triple treatment may serve as an efficacious delayed therapy, preventing brain insult propagation following sarin-induced refractory SE.
Assuntos
Lesões Encefálicas , Ketamina , Agentes Neurotóxicos , Estado Epiléptico , Ratos , Animais , Sarina/toxicidade , Agentes Neurotóxicos/toxicidade , Midazolam/farmacologia , Midazolam/uso terapêutico , Ratos Sprague-Dawley , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/tratamento farmacológico , Colinérgicos/efeitos adversos , Lesões Encefálicas/induzido quimicamenteRESUMO
OBJECTIVE: Given its key homeostatic role affecting mitochondria, ionotropic and metabotropic receptors, and voltage-gated ion channels, sigma-1 receptor (Sig1R) represents an interesting target for epilepsy management. Antiseizure effects of the positive allosteric modulator E1R have already been reported in acute seizure models. Although modulation of serotonergic neurotransmission is considered the main mechanism of action of fenfluramine, its interaction with Sig1R may be of additional relevance. METHODS: To further explore the potential of Sig1R as a target, we assessed the efficacy and tolerability of E1R and fenfluramine in two chronic mouse models, including an amygdala kindling paradigm and the intrahippocampal kainate model. The relative contribution of the interaction with Sig1R was analyzed using combination experiments with the Sig1R antagonist NE-100. RESULTS: Whereas E1R exerted pronounced dose-dependent antiseizure effects at well-tolerated doses in fully kindled mice, only limited effects were observed in response to fenfluramine, without a clear dose dependency. In the intrahippocampal kainate model, E1R failed to influence electrographic seizure activity. In contrast, fenfluramine significantly reduced the frequency of electrographic seizure events and their cumulative duration. Pretreatment with NE-100 reduced the effects of E1R and fenfluramine in the kindling model. Surprisingly, pre-exposure to NE-100 in the intrahippocampal kainate model rather enhanced and prolonged fenfluramine's antiseizure effects. SIGNIFICANCE: In conclusion, the kindling data further support Sig1R as an interesting target for novel antiseizure medications. However, it is necessary to further explore the preclinical profile of E1R in chronic epilepsy models with spontaneous seizures. Despite the rather limited effects in the kindling paradigm, the findings from the intrahippocampal kainate model suggest that it is of interest to further assess a possible broad-spectrum potential of fenfluramine.
Assuntos
Modelos Animais de Doenças , Epilepsia , Fenfluramina , Excitação Neurológica , Receptores sigma , Receptor Sigma-1 , Animais , Receptores sigma/antagonistas & inibidores , Receptores sigma/efeitos dos fármacos , Camundongos , Excitação Neurológica/efeitos dos fármacos , Fenfluramina/farmacologia , Epilepsia/tratamento farmacológico , Masculino , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Relação Dose-Resposta a Droga , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Tonsila do Cerebelo/efeitos dos fármacos , Tonsila do Cerebelo/fisiopatologia , Hipocampo/efeitos dos fármacos , Doença Crônica , Ácido Caínico/farmacologia , Camundongos Endogâmicos C57BLRESUMO
Epilepsy is a common neurological disorder, and the exploration of potential therapeutic drugs for its treatment is still ongoing. Vitamin D has emerged as a promising treatment due to its potential neuroprotective effects and anti-epileptic properties. This study aimed to investigate the effects of vitamin D on epilepsy and neuroinflammation in juvenile mice using network pharmacology and molecular docking, with a focus on the mammalian target of rapamycin (mTOR) signaling pathway. Experimental mouse models of epilepsy were established through intraperitoneal injection of pilocarpine, and in vitro injury models of hippocampal neurons were induced by glutamate (Glu) stimulation. The anti-epileptic effects of vitamin D were evaluated both in vivo and in vitro. Network pharmacology and molecular docking analysis were used to identify potential targets and regulatory pathways of vitamin D in epilepsy. The involvement of the mTOR signaling pathway in the regulation of mouse epilepsy by vitamin D was validated using rapamycin (RAPA). The levels of inflammatory cytokines (TNF-α, IL-1ß, and IL-6) were assessed by enzyme-linked immunosorbent assay (ELISA). Gene and protein expressions were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. The terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL) staining was used to analyze the apoptosis of hippocampal neurons. In in vivo experiments, vitamin D reduced the Racine scores of epileptic mice, prolonged the latency of epilepsy, and inhibited the production of TNF-α, IL-1ß, and IL-6 in the hippocampus. Furthermore, network pharmacology analysis identified RAF1 as a potential target of vitamin D in epilepsy, which was further confirmed by molecular docking analysis. Additionally, the mTOR signaling pathway was found to be involved in the regulation of mouse epilepsy by vitamin D. In in vitro experiments, Glu stimulation upregulated the expressions of RAF1 and LC3II/LC3I, inhibited mTOR phosphorylation, and induced neuronal apoptosis. Mechanistically, vitamin D activated the mTOR signaling pathway and alleviated mouse epilepsy via RAF1, while the use of the pathway inhibitor RAPA reversed this effect. Vitamin D alleviated epilepsy symptoms and neuroinflammation in juvenile mice by activating the mTOR signaling pathway via RAF1. These findings provided new insights into the molecular mechanisms underlying the anti-epileptic effects of vitamin D and further supported its use as an adjunctive therapy for existing anti-epileptic drugs.
Assuntos
Epilepsia , Simulação de Acoplamento Molecular , Farmacologia em Rede , Proteínas Proto-Oncogênicas c-raf , Transdução de Sinais , Serina-Treonina Quinases TOR , Vitamina D , Animais , Serina-Treonina Quinases TOR/metabolismo , Epilepsia/tratamento farmacológico , Epilepsia/metabolismo , Transdução de Sinais/efeitos dos fármacos , Camundongos , Vitamina D/farmacologia , Vitamina D/uso terapêutico , Masculino , Proteínas Proto-Oncogênicas c-raf/metabolismo , Doenças Neuroinflamatórias/tratamento farmacológico , Doenças Neuroinflamatórias/metabolismo , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Hipocampo/metabolismo , Hipocampo/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêuticoRESUMO
The ability of neural circuits to integrate information over time and across different cortical areas is believed an essential ingredient for information processing in the brain. Temporal and spatial correlations in cortex dynamics have independently been shown to capture these integration properties in task-dependent ways. A fundamental question remains if temporal and spatial integration properties are linked and what internal and external factors shape these correlations. Previous research on spatio-temporal correlations has been limited in duration and coverage, thus providing only an incomplete picture of their interdependence and variability. Here, we use long-term invasive EEG data to comprehensively map temporal and spatial correlations according to cortical topography, vigilance state and drug dependence over extended periods of time. We show that temporal and spatial correlations in cortical networks are intimately linked, decline under antiepileptic drug action, and break down during slow-wave sleep. Further, we report temporal correlations in human electrophysiology signals to increase with the functional hierarchy in cortex. Systematic investigation of a neural network model suggests that these dynamical features may arise when dynamics are poised near a critical point. Our results provide mechanistic and functional links between specific measurable changes in the network dynamics relevant for characterizing the brain's changing information processing capabilities.
Assuntos
Anticonvulsivantes , Vigília , Humanos , Anticonvulsivantes/farmacologia , Encéfalo/fisiologia , Mapeamento Encefálico/métodos , Imageamento por Ressonância Magnética/métodosRESUMO
γ-Aminobutyric acid (GABA) and GABA derivatives have attracted increased attention over the years in the fields of medicinal chemistry and chemical biology due to their interesting biological properties and synthetic relevance. Here, we report a short synthetic route to γ-(het)aryl- and γ-alkenyl-γ-aminobutyric acids, including the antiepileptic drug vigabatrin, from readily available donor-acceptor cyclopropanes and ammonia or methylamine. This protocol includes a facile synthesis of 2-oxopyrrolidine-3-carboxamides and their acid hydrolysis to γ-aryl- or γ-alkenyl-substituted GABAs, which can serve as perspective building blocks for the synthesis of various GABA-based N-heterocycles and bioactive compounds.
Assuntos
Vigabatrina , Ácido gama-Aminobutírico , Anticonvulsivantes/farmacologia , Vigabatrina/farmacologia , Pirrolidinas/química , Pirrolidinas/farmacologiaRESUMO
Natural compounds are increasingly being studied for their potential neuroprotective effects against inflammatory neurological diseases. Epilepsy is a common neurological disease associated with inflammatory processes, and around 30% of people with epilepsy do not respond to traditional treatments. Some flavonoids, when taken along with antiseizure medications can help reduce the likelihood of drug-resistant epilepsy. Baicalin, a plant-based compound, has been shown to possess pharmacological properties such as anti-inflammatory, neuroprotective, anticonvulsant, and antioxidant activities. In this study, we tested the effect of baicalin on an established model of pharmacologically induced seizure in zebrafish using measures of both locomotor behavior and calcium imaging of neuronal activity. The results of our study showed that, at the tested concentration, and contrary to other studies in rodents, baicalin did not have an anti-seizure effect in zebrafish larvae. However, given its known properties, other concentrations and approaches should be explored to determine if it could potentially have other beneficial effects, either alone or when administered in combination with classic antiseizure medications.
Assuntos
Cálcio , Flavonoides , Larva , Neurônios , Pentilenotetrazol , Convulsões , Peixe-Zebra , Animais , Flavonoides/farmacologia , Convulsões/tratamento farmacológico , Convulsões/induzido quimicamente , Larva/efeitos dos fármacos , Cálcio/metabolismo , Neurônios/efeitos dos fármacos , Modelos Animais de Doenças , Anticonvulsivantes/farmacologia , Relação Dose-Resposta a Droga , Convulsivantes/toxicidade , Locomoção/efeitos dos fármacos , Atividade Motora/efeitos dos fármacosRESUMO
Lamotrigine (LTG) is an antiepileptic drug with possible adverse effects on the female reproductive system. Curcumin was declared to improve ovarian performance. Therefore, this study aimed to clarify ovulatory dysfunction (OD) associated with LTG and the role of curcumin in ameliorating this dysfunction. Adult female Wister albino rats were assigned into four groups: negative control (received saline), positive control (received curcumin only), LTG, and LTG with curcumin groups. Drugs were administered for 90 days. The hormonal profile, including testosterone, estrogen, progesterone, luteinizing hormone, and follicle-stimulating hormone, in addition to the lipid profile and glycemic analysis, were tested. Oxidative stress biomarkers analysis in the ovaries and uterus and peroxisome proliferator-activated receptor-γ (PPAR-γ) gene expression were also included. Histopathological examination of ovarian and uterine tissues and immunohistochemical studies were also performed. Curcumin could improve the OD related to chronic LTG intake. That was proved by the normalization of the hormonal profile, glycemic control, lipidemic status, oxidative stress markers, and PPAR-γ gene expression. The histopathological and immunohistochemical examination of ovarian and uterine tissues revealed an improvement after curcumin administration. The results describe an obvious deterioration in ovarian performance with LTG through the effect on lipidemic status, PPAR-γ gene, and creating an oxidative stress condition in the ovaries of chronic users, with a prominent improvement with curcumin addition to the treatment protocol.
Assuntos
Curcumina , Ovário , Ratos , Feminino , Animais , Ovário/metabolismo , Curcumina/farmacologia , Lamotrigina/farmacologia , Anticonvulsivantes/farmacologia , Espécies Reativas de Oxigênio , PPAR gama/metabolismo , Ratos Wistar , Útero/metabolismoRESUMO
In this study, our goal was to synthesize novel aryl tacrine derivatives and assess their potential as anticancer, antibacterial agents, and enzyme inhibitors. We adopted a two-step approach, initiating with the synthesis of dibromotacrine derivatives 3 and 4 through the Friedlander reaction. These intermediates underwent further transformation into diarylated tacrine derivatives 3a-e and 4a-e using a Suzuki-Miyaura cross-coupling reaction. Thorough characterization of these novel diarylated tacrines was achieved using various spectroscopic techniques. Our findings highlighted the potent anticancer effects of these innovative compounds across a range of cancer cell lines, including lung, gynecologic, bone, colon, and breast cancers, while demonstrating low cytotoxicity against normal cells. Notably, these compounds surpassed the control drug, 5-Fluorouracil, in terms of antiproliferative activity in numerous cancer cell lines. Moreover, our investigation included an analysis of the inhibitory properties of these novel compounds against various microorganisms and cytosolic carbonic anhydrase enzymes. The results suggest their potential for further exploration as cancer-specific, enzyme inhibitory, and antibacterial therapeutic agents. Notably, four compounds, namely, 5,7-bis(4-(methylthio)phenyl)tacrine (3d), 5,7-bis(4-(trifluoromethoxy)phenyl)tacrine (3e), 2,4-bis(4-(trifluoromethoxy)phenyl)-7,8,9,10-tetrahydro-6H-cyclohepta[b]quinolin-11-amine (4e), and 6,8-dibromotacrine (3), emerged as the most promising candidates for preclinical studies.
Assuntos
Antineoplásicos , Neoplasias , Feminino , Humanos , Tacrina/farmacologia , Tacrina/química , Antifúngicos/farmacologia , Anticonvulsivantes/farmacologia , Antibacterianos/farmacologia , Antibacterianos/química , Inibidores Enzimáticos/farmacologia , Antineoplásicos/química , Relação Estrutura-Atividade , Estrutura MolecularRESUMO
BACKGROUND: Pyridine and its derivatives play a vital role in medicinal chemistry, serving as key scaffolds for drugs. The ability to bind to biological targets makes pyridine compounds significant, sparking interest in creating new pyridine-based drugs. Thus, the purpose of the research is to synthesize new thioalkyl derivatives of pyridine, predict their biological spectrum, study their psychotropic properties, and based on these findings, perform structure-activity relationships to assess pharmacophore functional groups. METHODS: Classical organic methods were employed for synthesizing new thioalkyl derivatives of pyridine, with a multifaceted pharmacological profiles. Various software packages and methods were employed to evaluate the biological spectrum of the newly synthesized compounds. For the evaluation of neurotropic activity of new synthesized compounds, some biological methods were used according to indicators characterizing anticonvulsant, sedative and antianxiety activity as well as side effects. RESULTS: Effective synthetic methods for 6-amino-4-phenyl-2-thio-2H-thiopyran-5-carboxylic acid ethyl ester, 2-amino substituted thiopyridine derivatives and 6-cycloamino-2-thioalkyl-4-phenylnicotinate derivatives were obtained in high yield. Predicted biological spectra and pharmacokinetic data indicated high gastrointestinal absorption and low blood-brain barrier passage for most compounds and demonstrated potential various biological effects, particularly psychotropic properties. Studied compounds demonstrated high anticonvulsant activity through antagonism with pentylenetetrazole. They exhibited low toxicity without inducing muscle relaxation in the studied doses. In psychotropic studies, the compounds displayed activating, sedative, and anxiolytic effects. Notably, the 6-amino-2-thioalkyl-4-phenylnicotinate derivatives demonstrated significant anxiolytic activity (about four times more compared to diazepam). They also exhibited pronounced sedative effects. Ethyl 2-({2-[(diphenylmethyl)amino]-2-oxoethyl}thio)-4-phenyl-6-pyrrolidin-1-ylnicotinate exhibited anxiolytic activity even two times greater than diazepam. Moreover, all studied compounds showed statistically significant antidepressant effects. Noteworthy ethyl 2-({2-oxo-2-[(tetrahydrofuran-2-ylmethyl)amino]ethyl}thio)-4-phenyl-6-pyrrolidin-1-ylnicotinate showcasing its unique psychotropic effect. CONCLUSIONS: The selected compounds demonstrate anticonvulsant properties, activating behavior, and anxiolytic effects, while simultaneously exhibiting antidepressant effects and these compounds as promising candidates for further exploration in the development of therapeutics with a broad spectrum of neuropsychiatric applications.
Assuntos
Ansiolíticos , Anticonvulsivantes , Piridinas , Relação Estrutura-Atividade , Piridinas/química , Piridinas/farmacologia , Piridinas/síntese química , Animais , Anticonvulsivantes/síntese química , Anticonvulsivantes/farmacologia , Anticonvulsivantes/química , Camundongos , Ansiolíticos/farmacologia , Ansiolíticos/síntese química , Ansiolíticos/química , Estrutura Molecular , Relação Dose-Resposta a Droga , Masculino , Convulsões/tratamento farmacológico , Hipnóticos e Sedativos/farmacologia , Hipnóticos e Sedativos/síntese química , Hipnóticos e Sedativos/química , PentilenotetrazolRESUMO
Based on the pharmacophore model of opioid receptors, our team recently synthesized a series of short-chain hemorphin peptide analogs containing non-natural amino acids. They demonstrated anticonvulsant and antinociceptive activity with low neurotoxicity. In the present study, a series of novel bioconjugates of N-modified hemorphin analogs containing second pharmacophore cinnamic acids (CA) or caffeic (KA) were synthesized by a traditional solid-phase Fmoc chemistry method for peptide synthesis. Electrochemical and fluorimetric analysis, in vivo anticonvulsant and antinociceptive activity in mice were conducted on the compounds. The three CA acid- (H4-CA, H5-CA, and H7-CA) and three KA acid- (H4-KA, H5-KA, and H7-KA) conjugated hemorphin derivatives exhibited potency at the highest doses of 2 µg/5 µl, administered by intracerebroventricular (icv) mode, against seizure spread in the maximal electroshock test (MES) in mice. The KA-conjugated H5-KA derivate, at the lowest dose, was the only compound that suppressed clonic seizures in the subcutaneous pentylenetetrazol (scPTZ) test. Except for the H5-CA, all tested CA acid- and KA acid-conjugated peptide derivates had the potency to increase the latency for clonic seizures in a dose-dependent mode. The activity against the psychomotor seizures in the 6-Hz test was detected only for the H4-CA (0.5 µg) and H4-KA (0.5 µg and 1 µg), respectively. All investigated peptides showed a more pronounced antinociceptive effect in the "intraplantar formalin" test compared to the "hot plate" test. Shorter chain analogs showed a better antinociceptive profile against tonic pain. The data suggest a DOR and KOR-mediated mechanism of action. According to the docking analysis, H7-CA showed a different antinociceptive profile than other investigated peptides. The novel peptide derivates did not exhibit neurotoxicity in the rotarod test. Our findings suggest that conjugated CA and KA morphine peptides can be used to develop novel morphine-related analogs with anticonvulsant and antinociceptive activity.