Repressor Element-1 Binding Transcription Factor (REST) as a Possible Epigenetic Regulator of Neurodegeneration and MicroRNA-Based Therapeutic Strategies.

Neurodegenerative disorders (NDD) have grabbed significant scientific consideration due to their fast increase in prevalence worldwide. The specific pathophysiology of the disease and the amazing changes in the brain that take place as it advances are still the top issues of contemporary research. Transcription factors play a decisive role in integrating various signal transduction pathways to ensure homeostasis. Disruptions in the regulation of transcription can result in various pathologies, including NDD. Numerous microRNAs and epigenetic transcription factors have emerged as candidates for determining the precise etiology of NDD. Consequently, understanding by what means transcription factors are regulated and how the deregulation of transcription factors contributes to neurological dysfunction is important to the therapeutic targeting of pathways that they modulate. RE1-silencing transcription factor (REST) also named neuron-restrictive silencer factor (NRSF) has been studied in the pathophysiology of NDD. REST was realized to be a part of a neuroprotective element with the ability to be tuned and influenced by numerous microRNAs, such as microRNAs 124, 132, and 9 implicated in NDD. This article looks at the role of REST and the influence of various microRNAs in controlling REST function in the progression of Alzheimer's disease (AD), Parkinson's disease (PD), and Huntington's disease (HD) disease. Furthermore, to therapeutically exploit the possibility of targeting various microRNAs, we bring forth an overview of drug-delivery systems to modulate the microRNAs regulating REST in NDD.

Phytotherapeutics against Alzheimer's Disease: Mechanism, Molecular Targets and Challenges for Drug Development.

Alzheimer's disease is inflating worldwide and is combatted by only a few approved drugs. At best, these drugs treat symptomatic conditions by targeting cholinesterase and N-methyl- D-aspartate receptors. Most of the clinical trials in progress are focused on developing disease-modifying agents that aim at single targets. The 'one drug-one target' approach is failing in the case of Alzheimer's disease due to its labyrinth etiopathogenesis. Traditional medicinal systems like Ayurveda use a holistic approach encompassing the legion of medicinal plants exhibiting multimodal activity. Recent advances in high-throughput technologies have catapulted the research in the arena of Ayurveda, specifically in identifying plants with potent anti-Alzheimer's disease properties and their phytochemical characterization. Nonetheless, clinical trials of very few herbal medicines are in progress. This review is a compendium of Indian plants and ayurvedic medicines against Alzheimer's disease and their paraphernalia. A record of 230 plants that are found in India with anti-Alzheimer's disease potential and about 500 phytochemicals from medicinal plants have been solicited with the hope of exploring the unexplored. Further, the molecular targets of phytochemicals isolated from commonly used medicinal plants, such as Acorus calamus, Bacopa monnieri, Convolvulus pluricaulis, Tinospora cordifolia and Withania somnifera, have been reviewed with respect to their multidimensional property, such as antioxidant, anti-inflammation, anti-aggregation, synaptic plasticity modulation, cognition, and memory-enhancing activity. In addition, the strengths and challenges in ayurvedic medicine that limit its use as mainstream therapy are discussed, and a framework for the development of herbal medicine has been proposed.

A Novel Five-Node Feed-Forward Loop Unravels miRNA-Gene-TF Regulatory Relationships in Ischemic Stroke.

The complex and interlinked cascade of events regulated by microRNAs (miRNAs), transcription factors (TF), and target genes highlight the multifactorial nature of ischemic stroke pathology. The complexity of ischemic stroke requires a wider assessment than the existing experimental research that deals with only a few regulatory components. Here, we assessed a massive set of genes, miRNAs, and transcription factors to build a miRNA-gene-transcription factor regulatory network to elucidate the underlying post-transcriptional mechanisms in ischemic stroke. Feed-forward loops (three-node, four-node, and novel five-node) were converged to establish regulatory relationships between miRNAs, TFs, and genes. The synergistic function of miRNAs in ischemic stroke was predicted and incorporated into a novel five-node feed-forward loop. Significant miRNA-TF pairs were identified using cumulative hypergeometric distribution. Two subnetworks were derived from the extensive miRNA-TF regulatory network and analyzed to predict the molecular mechanism relating the regulatory components. NFKB and STAT were identified to be the chief regulators of innate inflammatory and neuronal survival mechanisms, respectively. Exclusive novel interactions between miR-9 and miR-124 with TLX, BCL2, and HDAC4 were identified to explain the post-stroke induced neurogenesis mechanism. Therefore, this network-based approach to delineate miRNA, TF, and gene interactions might promote the development of effective therapeutics against ischemic stroke.

Amikacin Inhibits miR-497 Maturation and Exerts Post-ischemic Neuroprotection.

MicroRNAs (miRNAs) are a group of small non-coding RNAs that regulate numerous signaling pathways involved in cerebral ischemia reperfusion injury. Recent finding demonstrated that miR-497 promotes ischemic neuronal death by negatively regulating anti-apoptotic proteins and therefore serves as a promising therapeutic target for cerebral ischemic injury. In this study, we present a systematic computational approach that includes 3D modeling, docking-based virtual screening, and molecular dynamics simulation to identify small-molecule inhibitors of pre-miR-497 maturation. The top hit, aminoglycosidic antibiotic, amikacin, formed a stable complex with pre-miR-497. Later, the protective efficacy of amikacin was evaluated against oxygen-glucose deprivation (OGD) and reoxygenation-induced neuronal cell death in SH-SY5Y cells and mouse organotypic hippocampal slice cultures. To confirm the inhibitory potential of amikacin on miR-497 maturation, quantitative real-time PCR was performed to check the expression of bcl-2, one of the primary anti-apoptotic targets of miR-497. Additionally, the expression level of mature miR-497 was quantified using TaqMan® MiRNA Assay Kit. Amikacin treatment effectively reduced OGD-induced cell death compared to control groups both in vitro and organotypic hippocampal slice cultures. Further, amikacin effectively increased the expression of bcl-2 in SH-SY5Y cells subjected to OGD. Interestingly, SH-SY5Y cells treated with amikacin displayed decreased expression of miR-497, probably due to inhibition of pre-miRic form. Our study provides strong evidence that amikacin inhibits miR-497 maturation and promotes ischemic neuronal survival by upregulating anti-apoptotic protein, bcl-2. Future studies directed at evaluating the neuroprotective efficacy and mechanism of amikacin animal models may lead to new therapeutic opportunities for preventing neuronal death after stroke.

Novel RIPK3 inhibitors discovered through a structure-based approach exert post-ischemic neuroprotection.

Necroptosis or programmed necrosis is evident in various neurological disorders such as ischemic stroke. Receptor interacting serine/threonine protein kinase 3 (RIPK3) is one of the crucial targets of necroptosis and inhibition of this protein exerts neuroprotection. However, knowledge regarding the three-dimensional structure and binding site information of this protein is lacking. In the present study, structure-based in silico methods were implemented to identify the key amino acids in the RIPK3 binding site that might be responsible for ligand interactions. Further, novel RIPK3 inhibitors were identified through a dual ensemble screening strategy. Three inhibitors exhibited binding to RIPK3 in micromolar concentrations and exerted post-ischemic neuroprotection in vitro.

Ensembling and filtering: an effective and rapid in silico multitarget drug-design strategy to identify RIPK1 and RIPK3 inhibitors.

Necroptosis, a programmed necrosis pathway, is witnessed in diverse human diseases and is primarily regulated by receptor-interacting serine/threonine protein kinase 1 (RIPK1) and RIPK3. Ablation or inhibition of these individual proteins, or both, has been shown to be protective in various in vitro and in vivo disease models involving necroptosis. In this study, we propose an effective and rapid virtual screening strategy to identify multitarget inhibitors of both RIPK1 and RIPK3. It involves ensemble pharmacophore-based screening (EPS) of a compound database, post-EPS filtration (PEPSF) of the ligand hits, and multiple dockings. Structurally diverse inhibitors were identified through ensemble pharmacophore features, and the speed of this process was enhanced by filtering out the compounds containing cross-features. The stability of these inhibitors with both of the proteins was verified by means of molecular dynamics (MD) simulation. Graphical Abstract A generalized workflow employed in this study. Subsequent utilization of EPS and PEPSF might lead to reduced computational time and load.

Modelling the molecular mechanism of protein-protein interactions and their inhibition: CypD-p53 case study.

Cyclophilin D (CypD) is an important regulatory protein involved in mitochondrial membrane permeability transition and cell death. Further, the mitochondrial CypD-p53 axis is an important contributor to necroptosis, a form of programmed necrosis, involved in various cardiovascular and neurological disorders. The CypD ligand, Cyclosporin A (CsA), was identified as an inhibitor of this interaction. In this study, using computational methods, we have attempted to model the CypD-p53 interaction in order to delineate their mode of binding and also to disclose the molecular mechanism, by means of which CsA interferes with this interaction. It was observed that p53 binds at the CsA-binding site of CypD. The knowledge obtained from this modelling was employed to identify novel CypD inhibitors through structure-based methods. Further, the identified compounds were tested by a similar strategy, adopted during the modelling process. This strategy could be applied to study the mechanism of protein-protein interaction (PPI) inhibition and to identify novel PPI inhibitors.

Ensemble pharmacophore meets ensemble docking: a novel screening strategy for the identification of RIPK1 inhibitors.

Programmed cell death has been a fascinating area of research since it throws new challenges and questions in spite of the tremendous ongoing research in this field. Recently, necroptosis, a programmed form of necrotic cell death, has been implicated in many diseases including neurological disorders. Receptor interacting serine/threonine protein kinase 1 (RIPK1) is an important regulatory protein involved in the necroptosis and inhibition of this protein is essential to stop necroptotic process and eventually cell death. Current structure-based virtual screening methods involve a wide range of strategies and recently, considering the multiple protein structures for pharmacophore extraction has been emphasized as a way to improve the outcome. However, using the pharmacophoric information completely during docking is very important. Further, in such methods, using the appropriate protein structures for docking is desirable. If not, potential compound hits, obtained through pharmacophore-based screening, may not have correct ranks and scores after docking. Therefore, a comprehensive integration of different ensemble methods is essential, which may provide better virtual screening results. In this study, dual ensemble screening, a novel computational strategy was used to identify diverse and potent inhibitors against RIPK1. All the pharmacophore features present in the binding site were captured using both the apo and holo protein structures and an ensemble pharmacophore was built by combining these features. This ensemble pharmacophore was employed in pharmacophore-based screening of ZINC database. The compound hits, thus obtained, were subjected to ensemble docking. The leads acquired through docking were further validated through feature evaluation and molecular dynamics simulation.

Necroptosis: who knew there were so many interesting ways to die?

Conventional knowledge considered apoptosis as the sole form of programmed cell death during development, homeostasis and diseases, whereas necrosis was regarded as an unregulated and uncontrollable process. Recent revelations suggest that necrosis can also occur in a regulated, caspase-independent manner and shares characteristics with both necrosis and apoptosis. The major cell death processes namely apoptosis, autophagy and necrosis are interlinked and contain many common regulatory mechanisms. Mounting evidence indicates that necroptosis contributes to the pathogenesis of various diseases, including ischemic stroke, traumatic brain injury, neurodegenerative disorders and brain tumor. We present here an overview of the molecular mechanisms governing necroptosis and its connection with apoptosis and autophagy processes. Further, the necroptosis mechanisms underlying the neurodegeneration during ischemia reperfusion (I/R) injury are described, with an emphasis on the key proteins involved in this type of cell death. Knowledge regarding programmed cell death (PCD) with relevance to necroptosis may play a significant role in debilitating brain disorders.

A critical appraisal of the functional evolution of P2Y12 antagonists as antiplatelet drugs.

P2Y12 receptor mediated inhibition of platelet aggregation is one of the most explored and exploited pathways in antiplatelet drug therapy to prevent ischemic events in patients undergoing percutaneous coronary intervention (PCI) for the treatment of the acute coronary syndrome (ACS). Ticlopidine, Clopidogrel, Prasugrel, Ticagrelor, Cangrelor and Elinogrel are the P2Y12 inhibitors that act as antiplatelet drugs. In this review, the features of these drugs and the factors reported to be responsible for drug resistance or drug ineffectiveness were described. The features like drug metabolism, reversible or irreversible binding of drugs to their target protein and the mode of administration were observed to evolve along with the antiplatelet drugs. These features also include the drug-drug interactions, the pharmacogenetics and pharmacodynamics of P2Y12 inhibitors. We attempted to critically analyze how the desirable features were met by the P2Y12 inhibitors in the course of time. This review provides an overview of the evolution of P2Y12 inhibitors and may guide the researchers to develop better antiplatelet drugs in the future.