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Ubiquitination is a post-translational modification that elicits a variety of cellular responses. Deubiquitinases (DUBs) remove ubiquitin moieties from proteins and modulate cellular processes by counteracting the ubiquitin ligase activities. Ubiquitination and deubiquitination processes are tightly regulated by different mechanisms and their dysregulation is associated with many diseases. Discovery of DUB inhibitors could not only lead to therapeutics but also facilitate the understanding of ubiquitination/deubiquitination processes and their regulatory mechanisms. To enable the inhibitor discovery against DUBs, we developed a cell-based DUB assay that utilizes a cell-permeable ubiquitin probe, Biotin-cR 10 -Ub-PA, to covalently label DUBs in their native cellular environment. Amplified luminescent proximity homogeneous assay (Alpha, specifically AlphaLISA) is utilized to quantitatively assess the capture of the target DUB by the Biotin-cR 10 -Ub-PA probe. We demonstrated that this new cell-based DUB assay is robust and amenable to high-throughput screening. Human USP15 was selected as a DUB of interest and screened against a library of protease inhibitors as a proof of concept. In addition to the widely adopted pan-DUB inhibitor PR-619, several other DUB inhibitors from the library were also identified as hits. This new DUB assay can be readily adapted for inhibitor discovery against many other human DUBs to identify potent and cell-permeable inhibitors., Competing Interests: The authors declare no competing financial interest., (© 2024 American Chemical Society.)

Thrombosis, a key factor in most cardiovascular diseases, is a major contributor to human mortality. Existing antithrombotic agents carry a risk of bleeding. Consequently, there is a keen interest in discovering innovative antithrombotic agents that can prevent thrombosis without negatively impacting hemostasis. Platelets play crucial roles in both hemostasis and thrombosis. We have previously characterized calcium- and integrin-binding protein 1 (CIB1) as a key regulatory molecule that regulates platelet function. CIB1 interacts with several platelet proteins including integrin α IIb β 3 , the major glycoprotein receptor for fibrinogen on platelets. Given that CIB1 regulates platelet function through its interaction with α IIb β 3 , we developed a fluorescence polarization (FP) assay to screen for potential inhibitors. The assay was miniaturized to 1536-well and screened in quantitative high-throughput screening (qHTS) format against a diverse compound library of 14,782 compounds. After validation and selectivity testing using the FP assay, we identified 19 candidate inhibitors and validated them using an in-gel binding assay that monitors the interaction of CIB1 with α IIb cytoplasmic tail peptide, followed by testing of top hits by intrinsic tryptophan fluorescence (ITF) and microscale thermophoresis (MST) to ascertain their interaction with CIB1. Two of the validated hits shared similar chemical structures, suggesting a common mechanism of action. Docking studies further revealed promising interactions within the hydrophobic binding pocket of the target protein, particularly forming key hydrogen bonds with Ser180. The compounds exhibited a potent antiplatelet activity based on their inhibition of thrombin-induced human platelet aggregation, thus indicating that disruptors of the CIB1- α IIb β 3 interaction could carry a translational potential as antithrombotic agents., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

Computational models that predict pharmacokinetic properties are critical to deprioritize drug candidates that emerge as hits in high-throughput screening campaigns. We collected, curated, and integrated a database of compounds tested in 12 major end points comprising over 10,000 unique molecules. We then employed these data to build and validate binary quantitative structure-activity relationship (QSAR) models. All trained models achieved a correct classification rate above 0.60 and a positive predictive value above 0.50. To illustrate their utility in drug discovery, we used these models to predict the pharmacokinetic properties for drugs in the NCATS Inxight Drugs database. In addition, we employed the developed models to predict the pharmacokinetic properties of all compounds in the DrugBank. All models described in this paper have been integrated and made publicly available via the PhaKinPro Web-portal that can be accessed at https://phakinpro.mml.unc.edu/.

The theory based on numerical study of the system of hydrodynamic equations, which includes the director motion, shows that under the influence of crossed electric E and magnetic B fields, the director reorients in such a way that the transient quasiperiodic patterns may arise in microsized nematic volumes if the corresponding distortion mode has the fastest response and thus suppresses all other modes, including uniform ones. It has been shown that there is a threshold value of the amplitude of the thermal fluctuations of the director over the microsized nematic film which provides the nonuniform rotation mode rather than the uniform one, whereas the lower values of the amplitude dominate the uniform mode.

Globally, hepatocellular carcinoma (HCC) is one of the most commonly diagnosed cancers and a leading cause of cancer-related death. We previously identified an immune evasion pathway whereby tumor cells produce retinoic acid (RA) to promote differentiation of intratumoral monocytes into protumor macrophages. Retinaldehyde dehydrogenase 1 (RALDH1), RALDH2, and RALDH3 are the three isozymes that catalyze RA biosynthesis. In this study, we have identified RALDH1 as the key driver of RA production in HCC and demonstrated the efficacy of RALDH1-selective inhibitors (Raldh1-INH) in suppressing RA production by HCC cells. Raldh1-INH restrained tumor growth in multiple mouse models of HCC by reducing the number and tumor-supporting functions of intratumoral macrophages as well as increasing T-cell infiltration and activation within tumors. Raldh1-INH also displayed favorable pharmacokinetic, pharmacodynamic, and toxicity profiles in mice thereby establishing them as promising new drug candidates for HCC immunotherapy., (©2023 American Association for Cancer Research.)

The bidirectional relationship between cerebral structures and the gastrointestinal tract involving the microbiota embraces the scientific concept of the microbiota-gut-brain axis. The gut microbiome plays an important role in many physiological and biochemical processes of the human body, in the immune response and maintenance of homeostasis, as well as in the regulation of circadian rhythms. There is a relationship between the higher prevalence of a number of neurological disorders, sleep disorders and changes in the intestinal microbiota, which actualizes the study of the complex mechanisms of such correlation for the development of new treatment and prevention strategies. Environmental factors associated with excessive light exposure can aggravate the gut dysbiosis of intestinal microflora, and as a result, lead to sleep disturbances. This review examines the integrative mechanisms of sleep regulation associated with the gut microbiota (the role of neurotransmitters, short-chain fatty acids, unconjugated bile acids, bacterial cell wall components, cytokines). Taking into account the influence of gut dysbiosis as a risk factor in the development of various diseases, the authors systematize key aspects and modern scientific data on the importance of microflora balance to ensure optimal interaction along the microbiota-gut-brain axis in the context of the regulatory role of the sleep-wake cycle and its disorders.

Computerized cognitive training (CCT) is a rapidly developing area of cognitive rehabilitation at the intersection of information technology and healthcare, the constantly updated results of which can be successfully translated into practical application in clinical medicine and in particular in gerontology. The basis of CCT, as a non-invasive method of influencing the functional activity of the brain and the processes of neuroplasticity, is software for stimulating cognitive functions in order to improve their productivity. The level of scientific and practical interest in CCP technology is growing rapidly. The article reports on the current state of research on the use of CCT aimed at correcting cognitive impairment. The purpose of this work is to systematize the available scientific data in this area, as well as to promote further integration of research in the field of information technology into clinical practice, in particular, to study the potential of CCT as a promising therapeutic tool in the paradigm of successful aging and prevention of the progression of cognitive impairment. This noninvasive intervention may improve global cognitive function in patients with clinically defined impairments and during normal aging in cognitively healthy older adults. However, new studies with fully comparable protocols are needed to evaluate in more detail the duration of the effect and the effectiveness of CCT in preventing cognitive decline in the long term.

Molecular dynamics simulations were performed to study structural and dynamic properties of polar butanamine/water/butanamine, pentanoic acid/water/pentanoic acid, butanethiol/water/butanethiol, and nonpolar pentane/water/pentane systems. The mass density profiles along the interface normal to the organic liquid/water system, the difference in the local structure of H 2 O molecules in bulk and in the vicinity of interface, as well as the diffusion behavior of water molecules at the interface with above-mentioned organic liquids have been investigated. Our MD simulation has shown that the diffusion of water molecules across the water/organic liquid interface is influenced by the hydrogen bonds n HB between water molecules and the terminal groups of organic liquids. It was found that the loss of the hydrogen bonds n HB in the nonpolar organic liquid leads to a decrease in the value of the normal component of the diffusion coefficient D z , while the tangential diffusion coefficients, both D x and D y , increase., (© 2023 Wiley Periodicals LLC.)

Hits from high-throughput screening (HTS) of chemical libraries are often false positives due to their interference with assay detection technology. In response, we generated the largest publicly available library of chemical liabilities and developed "Liability Predictor," a free web tool to predict HTS artifacts. More specifically, we generated, curated, and integrated HTS data sets for thiol reactivity, redox activity, and luciferase (firefly and nano) activity and developed and validated quantitative structure-interference relationship (QSIR) models to predict these nuisance behaviors. The resulting models showed 58-78% external balanced accuracy for 256 external compounds per assay. QSIR models developed and validated herein identify nuisance compounds among experimental hits more reliably than do popular PAINS filters. Both the models and the curated data sets were implemented in "Liability Predictor," publicly available at https://liability.mml.unc.edu/. "Liability Predictor" may be used as part of chemical library design or for triaging HTS hits.

Electrically driven kinklike distortion regimes in a microsized liquid crystal channel have been investigated both experimentally and analytically. Kinklike distortion waves were excited by the interaction between the electric field E and the gradient ∇n[over ̂] of the director field in a homogeneously aligned liquid crystal (HALC) channel. Having obtained the evolution of the normalized light intensity, which was recorded by the high-speed camera, the process of excitation and evolution of the traveling wave in the HALC channel was visualized for the first time. It was shown, based on a nonlinear extension of the classical Ericksen-Leslie theory, that in the case when the electric field E≫E_{th}, the flow of liquid crystal material completely stops and a new mechanism for converting the electric field arises in the form of the electrically driven distorting traveling kinklike wave, which can be excited in the LC channel, composed of 4-n-pentyl-4^{'}-cyanobiphenyl molecules.

Diseases caused by new viruses cost thousands if not millions of human lives and trillions of dollars. We have identified, collected, curated, and integrated all chemogenomics data from ChEMBL for 13 emerging viruses that hold the greatest potential threat to global human health. By identifying and solving several challenges related to data annotation accuracy, we developed a highly curated and thoroughly annotated database of compounds tested in both phenotypic and target-based assays for these viruses that we dubbed SMACC (Small Molecule Antiviral Compound Collection). The pilot version of the SMACC database contains over 32,500 entries for 13 viruses. By analyzing data in SMACC, we have identified ∼50 compounds with polyviral inhibition profile, mostly covering flavi- and coronaviruses. The SMACC database may serve as a reference for virologists and medicinal chemists working on the development of novel BSA agents in preparation for future viral outbreaks. SMACC is publicly available at https://smacc.mml.unc.edu., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: AT and ENM are co-founders of Predictive, LLC, which develops computational methodologies and software for toxicity prediction. All other authors declare they have nothing to disclose., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Programmed ferroptotic death eliminates cells in all major organs and tissues with imbalanced redox metabolism due to overwhelming iron-catalyzed lipid peroxidation under insufficient control by thiols (Glutathione (GSH)). Ferroptosis has been associated with the pathogenesis of major chronic degenerative diseases and acute injuries of the brain, cardiovascular system, liver, kidneys, and other organs, and its manipulation offers a promising new strategy for anticancer therapy. This explains the high interest in designing new small-molecule-specific inhibitors against ferroptosis. Given the role of 15-lipoxygenase (15LOX) association with phosphatidylethanolamine (PE)-binding protein 1 (PEBP1) in initiating ferroptosis-specific peroxidation of polyunsaturated PE, we propose a strategy of discovering antiferroptotic agents as inhibitors of the 15LOX/PEBP1 catalytic complex rather than 15LOX alone. Here we designed, synthesized, and tested a customized library of 26 compounds using biochemical, molecular, and cell biology models along with redox lipidomic and computational analyses. We selected two lead compounds, FerroLOXIN-1 and 2, which effectively suppressed ferroptosis in vitro and in vivo without affecting the biosynthesis of pro-/anti-inflammatory lipid mediators in vivo. The effectiveness of these lead compounds is not due to radical scavenging or iron-chelation but results from their specific mechanisms of interaction with the 15LOX-2/PEBP1 complex, which either alters the binding pose of the substrate [eicosatetraenoyl-PE (ETE-PE)] in a nonproductive way or blocks the predominant oxygen channel thus preventing the catalysis of ETE-PE peroxidation. Our successful strategy may be adapted to the design of additional chemical libraries to reveal new ferroptosis-targeting therapeutic modalities.

Amyotrophic lateral sclerosis (ALS) is manifested as skeletal muscle denervation, loss of motor neurons and finally severe respiratory failure. Mutations of RNA-binding protein FUS are one of the common genetic reasons of ALS accompanied by a 'dying back' type of degeneration. Using fluorescent approaches and microelectrode recordings, the early structural and functional alterations in diaphragm neuromuscular junctions (NMJs) were studied in mutant FUS mice at the pre-onset stage. Lipid peroxidation and decreased staining with a lipid raft marker were found in the mutant mice. Despite the preservation of the end-plate structure, immunolabeling revealed an increase in levels of presynaptic proteins, SNAP-25 and synapsin 1. The latter can restrain Ca 2+ -dependent synaptic vesicle mobilization. Indeed, neurotransmitter release upon intense nerve stimulation and its recovery after tetanus and compensatory synaptic vesicle endocytosis were markedly depressed in FUS mice. There was a trend to attenuation of axonal [Ca 2+ ] in increase upon nerve stimulation at 20 Hz. However, no changes in neurotransmitter release and the intraterminal Ca 2+ transient in response to low frequency stimulation or in quantal content and the synchrony of neurotransmitter release at low levels of external Ca 2+ were detected. At a later stage, shrinking and fragmentation of end plates together with a decrease in presynaptic protein expression and disturbance of the neurotransmitter release timing occurred. Overall, suppression of synaptic vesicle exo-endocytosis upon intense activity probably due to alterations in membrane properties, synapsin 1 levels and Ca 2+ kinetics could be an early sign of nascent NMJ pathology, which leads to neuromuscular contact disorganization.

Dietary supplements and natural products are often marketed as safe and effective alternatives to conventional drugs, but their safety and efficacy are not well regulated. To address the lack of scientific data in these areas, we assembled a collection of Dietary Supplements and Natural Products (DSNP), as well as Traditional Chinese Medicinal (TCM) plant extracts. These collections were then profiled in a series of in vitro high-throughput screening assays, including a liver cytochrome p450 enzyme panel, CAR/PXR signaling pathways, and P-glycoprotein (P-gp) transporter assay activities. This pipeline facilitated the interrogation of natural product-drug interaction (NaPDI) through prominent metabolizing pathways. In addition, we compared the activity profiles of the DSNP/TCM substances with those of an approved drug collection (the NCATS Pharmaceutical Collection or NPC). Many of the approved drugs have well-annotated mechanisms of action (MOAs), while the MOAs for most of the DSNP and TCM samples remain unknown. Based on the premise that compounds with similar activity profiles tend to share similar targets or MOA, we clustered the library activity profiles to identify overlap with the NPC to predict the MOAs of the DSNP/TCM substances. Our results suggest that many of these substances may have significant bioactivity and potential toxicity, and they provide a starting point for further research on their clinical relevance., Competing Interests: The authors declare no competing financial interest., (Not subject to U.S. Copyright. Published 2023 by American Chemical Society.)

A skeletal photorearrangement involving UV-induced 6π-electrocyclization of diarylethenes with various ethene bridges has been studied. It has been found that deprotonation is the predominant step among the three possible alternative reaction pathways (radical abstraction, deprotonation, or sigmatropic shift) following 6π-electrocyclization, and incorporation of an electronegative carbonyl group into the geminal position to the phenyl residue results in a reduction in the reaction time and an increase in the yield of the desired product. The significant increase in the reaction time in less polar solvents (toluene, TCM) also indicates a large contribution of the deprotonation step to the skeletal photorearrangement of diarylethenes. Performing the reaction in toluene in the presence of tertiary amines leads to a reduction in the reaction time and an increase in the yield of the desired product. The best results were achieved when the reaction was carried out in toluene in the presence of DIPEA. The experimental results are in good agreement with the DFT calculations.

Objective: To evaluate the modifiable daily behavior patterns associated with increased anxiety indicators in the general population in response to the COVID-19 pandemic., Material and Methods: The study examined the characteristics of the Russian population ( n= 7777) of the international multicenter project COMET-G. In particular, variables were targeted to describe deviations in the behavior of adults during the period of application of measures of social isolation in connection with the pandemic, and revealing a relationship with the total score on the Spielberger State Anxiety Scale (STAI-S). Among these variables, experts selected those that could potentially be subject to change in the short term, that is, act as manageable or modifiable risk factors for the development of anxiety. The selected variables were analyzed in a statistical PLS-model to identify indicators that make the most significant contribution to the increase in the total anxiety score., Results: Our statistical model explained 48.4% of the variability in the STAI-S anxiety total scores related to changes in daily life habits. In particular, doom-scrolling/doom-surfing about the spread of the virus and the COVID-19 pandemic, changes in sleep patterns and usual daily life activities due to social isolation measures presented as factors significantly contributing to the increase of state anxiety., Conclusion: Given the manageable or modifiable risk factors that we have identified, public awareness and therapeutic recommendations, pointing to the need to (I) control the amount of time spent in the internet and monitor their internet-based content consumption, (II) regulate sleep-wake patterns, (III) maintain daily habits and household activities, may reduce the likelihood of developing anxiety disorders in the context of the impact of a global chronic stress due to the COVID-19 pandemic and associated social isolation measures.

Selective and progressive death of neurons is a characteristic feature of the process of neurodegeneration and leads to corresponding neuronal dysfunctions. Neurodegenerative diseases represent a heterogeneous group of clinically distinct disorders with similar molecular mechanisms of pathogenesis. They are based on the processes of abnormal aggregation of proteins, the formation of fibrillary insoluble structures and their deposition in the form of histopathological inclusions in the tissues of the nervous system. Disturbance of homeostatic functions that regulate neuronal ion and energy metabolism, biosynthesis and degradation of proteins and nucleotides, chronic hypoxia and the penetration of toxic and inflammatory substances into the brain from the bloodstream not only cause metabolic changes associated with age and disorders in the sleep-wake cycle, but also contribute to the development of neurodegenerative diseases. In animal studies, clearance pathways have been identified in which solutes and specific tracers are excreted perivascular into the meningeal lymphatics. The glymphatic pathway promotes the removal of metabolites, including Aβ amyloid and tau protein, from the parenchymal extracellular space of the brain. The glymphatic system is discussed to be more efficient during natural sleep, and fluid dynamics through this pathway exhibit daily fluctuations and are under circadian control. This review systematizes the key aspects and the data of recent research on the role of the glymphatic pathway and astroglial AQP-4 as its main determinant in maintaining homeostatic fluid circulation in the brain in normal and pathological conditions, in particular in relation to the regulatory role of the sleep-wake cycle and in development of neurodegeneration.

Selective and progressive death of neurons is a characteristic feature of the process of neurodegeneration and leads to corresponding neuronal dysfunctions. Neurodegenerative diseases represent a heterogeneous group of clinically distinct disorders with similar molecular mechanisms of pathogenesis. They are based on the processes of abnormal aggregation of proteins, the formation of fibrillary insoluble structures and their deposition in the form of histopathological inclusions in the tissues of the nervous system. Disturbance of homeostatic functions that regulate neuronal ion and energy metabolism, biosynthesis and degradation of proteins and nucleotides, chronic hypoxia and the penetration of toxic and inflammatory substances into the brain from the bloodstream not only cause metabolic changes associated with age and disorders in the sleep-wake cycle, but also contribute to the development of neurodegenerative diseases. In animal studies, clearance pathways have been identified in which solutes and specific tracers are excreted perivascular into the meningeal lymphatics. The glymphatic pathway promotes the removal of metabolites, including Aβ amyloid and tau protein, from the parenchymal extracellular space of the brain. The glymphatic system is discussed to be more efficient during natural sleep, and fluid dynamics through this pathway exhibit daily fluctuations and are under circadian control. This review systematizes the key aspects and scientific data of recent studies on the role of the glymphatic pathway and astroglial AQP-4 as its main determinant in maintaining homeostatic fluid circulation in the brain in normal and pathological conditions, in particular in relation to the regulatory role of the sleep-wake cycle and in development of neurodegeneration.

Chitosan-decorated liposomes were proposed for the first time for the intranasal delivery of acetylcholinesterase (AChE) reactivator pralidoxime chloride (2-PAM) to the brain as a therapy for organophosphorus compounds (OPs) poisoning. Firstly, the chitosome composition based on phospholipids, cholesterol, chitosans (Cs) of different molecular weights, and its arginine derivative was developed and optimized. The use of the polymer modification led to an increase in the encapsulation efficiency toward rhodamine B (RhB; ~85%) and 2-PAM (~60%) by 20% compared to conventional liposomes. The formation of monodispersed and stable nanosized particles with a hydrodynamic diameter of up to 130 nm was shown using dynamic light scattering. The addition of the polymers recharged the liposome surface (from -15 mV to +20 mV), which demonstrates the successful deposition of Cs on the vesicles. In vitro spectrophotometric analysis showed a slow release of substrates (RhB and 2-PAM) from the nanocontainers, while the concentration and Cs type did not significantly affect the chitosome permeability. Flow cytometry and fluorescence microscopy qualitatively and quantitatively demonstrated the penetration of the developed chitosomes into normal Chang liver and M-HeLa cervical cancer cells. At the final stage, the ability of the formulated 2-PAM to reactivate brain AChE was assessed in a model of paraoxon-induced poisoning in an in vivo test. Intranasal administration of 2-PAM-containing chitosomes allows it to reach the degree of enzyme reactivation up to 35 ± 4%.

Differential scanning fluorimetry is a rapid and economical biophysical technique used to monitor perturbations to protein structure during a thermal gradient, most often by detecting protein unfolding events through an environment-sensitive fluorophore. By employing an NTA-complexed fluorophore that is sensitive to nearby structural changes in histidine-tagged protein, a robust and sensitive differential scanning fluorimetry (DSF) assay is established with the specificity of an affinity tag-based system. We developed, optimized, and miniaturized this HIS-tag DSF assay (HIS-DSF) into a 1536-well high-throughput biophysical platform using the Borrelial high temperature requirement A protease (BbHtrA) as a proof of concept for the workflow. A production run of the BbHtrA HIS-DSF assay showed a tight negative control group distribution of T m values with an average coefficient of variation of 0.51% and median coefficient of variation of compound T m of 0.26%. The HIS-DSF platform will provide an additional assay platform for future drug discovery campaigns with applications in buffer screening and optimization, target engagement screening, and other biophysical assay efforts., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Ronzetti, Baljinnyam, Itkin, Jain, Rai, Zakharov, Pal and Simeonov.)

Cancer is one of the most serious health problems that usually require heavy medical treatment. It is important to ensure that no additional burden is placed on patients due to the modes of administration and/or poor quality of pharmaceuticals. In this regard, understanding, quantifying, and improving the photostability (resistance to UV light or sunlight) of drugs is among the important elements that can improve the patient's quality of life. In this work, the photochemical properties of a wide range of furanone analogues of combretastatin A-4 and their antiproliferative activity against A-431 epidermoid carcinoma cells were studied in a search for compounds with improved photostability and antiproliferative activity. It was found that the incorporation of an arylidene moiety led to a significant improvement in photostability, while the antiproliferative activity strongly depends on the nature of the aryl residue in the arylidene moiety. The high photostability of arylidenes was achieved due to the delocalization of the central double bond of the 1,3,5-hexatriene system, which limited the 6π-electrocyclization. The best results in terms of antiproliferative activity were obtained for thiophene arylidene (IC 50 = 0.6 μM) and 3,4-diarylfuran (IC 50 = 0.047 μM). The obtained results address the lack of data available now in scientific literature on the photodegradation of combretastatin A-4 analogues and should be taken into account in studies of the side effects of pharmaceuticals based on them.

The purpose of this article is to describe the physical mechanism responsible for the appearance of both traveling and nontraveling distortions in a microsized homogeneously aligned nematic (HAN) film under the effect of a large electric field. Numerical studies have been carried out to describe both the traveling and nontraveling dynamic reorientation of the director's field in a thin, in a few tens of micrometers, the HAN film under the effect of a large electric field E (∼1.0V/μm). It is shown that in response to the electric field E applied parallel to the bounding surfaces, the torques acting on the director n[over ̂] may excite the traveling distortion wave propagating normally to both boundaries, whose resemblance to a kinklike wave increases with increasing applied electric field E. Calculations show that in the HAN film the physical mechanism that is responsible for the electric-field-induced distortion of the director field n[over ̂] in the form of traveling wave provides a much faster relaxation regime than in the case of the nontraveling mode.

Nucleic acid nanoparticles, or NANPs, rationally designed to communicate with the human immune system, can offer innovative therapeutic strategies to overcome the limitations of traditional nucleic acid therapies. Each set of NANPs is unique in their architectural parameters and physicochemical properties, which together with the type of delivery vehicles determine the kind and the magnitude of their immune response. Currently, there are no predictive tools that would reliably guide the design of NANPs to the desired immunological outcome, a step crucial for the success of personalized therapies. Through a systematic approach investigating physicochemical and immunological profiles of a comprehensive panel of various NANPs, the research team developes and experimentally validates a computational model based on the transformer architecture able to predict the immune activities of NANPs. It is anticipated that the freely accessible computational tool that is called an "artificial immune cell," or AI-cell, will aid in addressing the current critical public health challenges related to safety criteria of nucleic acid therapies in a timely manner and promote the development of novel biomedical tools., (© 2022 The Authors. Small published by Wiley-VCH GmbH.)

Venglustat is a known allosteric inhibitor for ceramide glycosyltransferase, investigated in diseases caused by lysosomal dysfunction. Here, we identified venglustat as a potent inhibitor (IC 50 = 0.42 μM) of protein N-terminal methyltransferase 1 (NTMT1) by screening 58,130 compounds. Furthermore, venglustat exhibited selectivity for NTMT1 over 36 other methyltransferases. The crystal structure of NTMT1-venglustat and inhibition mechanism revealed that venglustat competitively binds at the peptide substrate site. Meanwhile, venglustat potently inhibited protein N-terminal methylation levels in cells (IC 50 = 0.5 μM). Preliminary structure-activity relationships indicated that the quinuclidine and fluorophenyl parts of venglustat are important for NTMT1 inhibition. In summary, we confirmed that venglustat is a bona fide NTMT1 inhibitor, which would advance the study on the biological roles of NTMT1. Additionally, this is the first disclosure of NTMT1 as a new molecular target of venglustat, which would cast light on its mechanism of action to guide the clinical investigations.

Here, we establish different ways of light-triggered droplet manipulation such as reversible wetting, splitting, merging, and transport. The unique features of our approach are that the changes in the wetting properties of microscopic droplets of isotropic (oil) or anisotropic (liquid crystalline) liquids adsorbed on photoswitchable films can be triggered just by application of soft optical stimuli, which lead to dynamical, reversible changes in the local morphology of the structured surfaces. The adaptive films consist of an azobenzene-containing surfactant ionically attached to oppositely charged polymer chains. Under exposure to irradiation with light, the azobenzene photoisomerizes between two states, nonpolar trans -isomer and polar cis -isomer, resulting in the corresponding changes in the surface energy and orientation of the surfactant tails at the interface. Additionally, the local increase in the surface temperature due to absorption of light by the azobenzene groups enables diverse processes of manipulation of the adsorbed small droplets, such as the reversible increase of the droplet basal area up to 5 times, anisotropic wetting during irradiation with modulated light, and precise partition of the droplet into many small pieces, which can then be merged on demand to the desired number of larger droplets. Moreover, using a moving focused light spot, we experimentally demonstrate and theoretically explain the locomotion of the droplet over macroscopic distances with a velocity of up to 150 μm·s -1 . Our findings could lead to the ultimate application of a programmable workbench for manipulating and operating an ensemble of droplets, just using simple and gentle optical stimuli.

An interface between a dielectric and a medium containing charge carriers with moderate mobility is considered. In equilibrium, stochastic fluxes of positive and negative particles toward the surface have equal average current density j 0 , and we suppose that the surface absorbs all falling charges. All over the surface, this results in the emergence of oppositely charged spots of various sizes D fluctuating and interacting with each other. Fourier expansion reduces this collection of interacting spots to the ensemble of independently fluctuating charge density waves. An exact solution of the Poisson equation for a single wave on a flat surface was obtained and provided strict proof that a fluctuating electric field is quite strong just above each charge spot but diminishes exponentially with the distance from the plane. The lifetime τ of a charge spot is inversely proportional to the density j 0 of the stochastic current while proportional to j 0 τ fluctuation's amplitudes independent of j 0 . The fluctuation's parameter dependence on the charge spot's size D can vary according to the conducting medium properties.

The COVID-19 pandemic has had enormous health, economic, and social consequences. Vaccines have been successful in reducing rates of infection and hospitalization, but there is still a need for acute treatment of the disease. We investigate whether compounds that bind the human angiotensin-converting enzyme 2 (ACE2) protein can decrease SARS-CoV-2 replication without impacting ACE2's natural enzymatic function. Initial screening of a diversity library resulted in hit compounds active in an ACE2-binding assay, which showed little inhibition of ACE2 enzymatic activity (116 actives, success rate ∼4%), suggesting they were allosteric binders. Subsequent application of in silico techniques boosted success rates to ∼14% and resulted in 73 novel confirmed ACE2 binders with K d values as low as 6 nM. A subsequent SARS-CoV-2 assay revealed that five of these compounds inhibit the viral life cycle in human cells. Further effort is required to completely elucidate the antiviral mechanism of these ACE2-binders, but they present a valuable starting point for both the development of acute treatments for COVID-19 and research into the host-directed therapy., Competing Interests: The authors declare the following competing financial interest(s): A.T. and E.N.M. are co-founders of Predictive, LLC, which develops computational methodologies and software for toxicity prediction., (© 2022 American Chemical Society.)

The purpose of this article is to describe the physical mechanism responsible for the appearance of both spatially periodic and kinklike distortions in a homogeneously aligned microsized nematic volume under the effect of crossed electric and magnetic fields. Numerical studies were carried out to describe the dynamic reorientation of the director^{'}s field in a thick liquid crystal (LC) cell (∼200μm) under the effect of a large electric field E (∼1.0V/μm) directed at an angle α close to a right angle to magnetic field B (∼7.0T). It is shown that under the effect of E directed at α∼89.96^{∘} to B, at least two scenarios of reorientation of the director field can be realized. First, in response to the suddenly applied electric field, spatially periodic patterns can appear in an initially uniformly aligned nematic domain. Second, when the same crossed external fields are applied at the smaller angle α∼88.81^{∘} to each other, the mode of uniform director reorientation is dominated. In the case when the electric field E≫E_{th} is applied orthogonally to both horizontal bounding surfaces of the LC cell and the magnetic field is turned off, in the microsized nematic volume the distortion in the form of the kinklike wave spreading normally to the horizontal bounding surfaces with the velocity in a few meters per second can be excited.

The global health emergency posed by the outbreak of Zika virus (ZIKV), an arthropod-borne flavivirus causing severe neonatal neurological conditions, has subsided, but there continues to be transmission of ZIKV in endemic regions. As such, there is still a medical need for discovering and developing therapeutical interventions against ZIKV. To identify small-molecule compounds that inhibit ZIKV disease and transmission, we screened multiple small-molecule collections, mostly derived from natural products, for their ability to inhibit wild-type ZIKV. As a primary high-throughput screen, we used a viral cytopathic effect (CPE) inhibition assay conducted in Vero cells that was optimized and miniaturized to a 1536-well format. Suitably active compounds identified from the primary screen were tested in a panel of orthogonal assays using recombinant Zika viruses, including a ZIKV Renilla luciferase reporter assay and a ZIKV mCherry reporter system. Compounds that were active in the wild-type ZIKV inhibition and ZIKV reporter assays were further evaluated for their inhibitory effects against other flaviviruses. Lastly, we demonstrated that wild-type ZIKV is able to infect a 3D-bioprinted outer-blood-retina barrier tissue model and disrupt its barrier function, as measured by electrical resistance. One of the identified compounds (3-Acetyl-13-deoxyphomenone, NCGC00380955) was able to prevent the pathological effects of the viral infection on this clinically relevant ZIKV infection model., Competing Interests: The authors have declared that no competing interests exist.

A molecular model describing the effective anchoring energy of a liquid crystal (LC) system, composed of 4-n-pentyl-4^{'}-cyanobiphenyl (5CB) molecules deposited on a photosensitive azobenzene layer consisting of 6Az10PVA molecules, is proposed. This model takes into account the interaction between the surface polarization in the LC phase and the surface electric field, arising from the surface charge density. Within the framework of this molecular model, the mechanism responsible for the anchoring transition in the LC phase from homeotropic to planar alignment and vice versa, caused by trans-cis and cis-trans conformational changes in the monolayer 6Az10PVA after laser radiation, is described. It is shown that using experimental data for the voltage across the 6Az10PVA+5CB film, obtained by the surface potential technique, the charge separation during the conformational changing, caused by the laser irradiation, may lead to an anchoring transition induced by trans-cis-trans isomerization in the photosensitive azobenzene monolayer. The calculated values of the isothermal surface pressure diagram π-A showed that the surface area A per LC molecule is noticeably higher for the cis configuration, which reflects a less efficient packaging than in the trans state.

The skeletal photorearrangement including 6π-electrocyclization induced by UV light of ortho -halogen-substituted diarylethenes has been studied. It has been found that the reaction pathways leading to bi- or tricyclic frameworks depend on the kind of halogen substituent and solvent. Photocyclization with halogen abstraction leads to bicyclic fused aromatics, while the tricyclic frameworks are formed due to the tandem 6π-electrocyclization/sigmatropic shift reaction. THF is preferred as the solvent in the former process and chloroform in the latter reaction. It was found for the first time that, owing to the ability of this series of diarylethenes to undergo skeletal photorearrangement with the release of the bromide cation, they can be used both as brominating agents and as Lewis acids for catalyzing electrophilic reactions.

Current high-throughput screening assay optimization is often a manual and time-consuming process, even when utilizing design-of-experiment approaches. A cross-platform, Cloud-based Bayesian optimization-based algorithm was developed as part of the National Center for Advancing Translational Sciences (NCATS) ASPIRE (A Specialized Platform for Innovative Research Exploration) Initiative to accelerate preclinical drug discovery. A cell-free assay for papain enzymatic activity was used as proof of concept for biological assay development and system operationalization. Compared with a brute-force approach that sequentially tested all 294 assay conditions to find the global optimum, the Bayesian optimization algorithm could find suitable conditions for optimal assay performance by testing 21 assay conditions on average, with up to 20 conditions being tested simultaneously, as confirmed by repeated simulation. The algorithm could achieve a sevenfold reduction in costs for lab supplies and high-throughput experimentation runtime, all while being controlled from a remote site through a secure connection. Based on this proof of concept, this technology is expected to be applied to more complex biological assays and automated chemistry reaction screening at NCATS, and should be transferable to other institutions.

The estrogen receptor α (ERα) is a target of intense pharmacological intervention and toxicological biomonitoring. Current methods to directly quantify cellular levels of ERα involve antibody-based assays, which are labor-intensive and of limited throughput. In this study, we generated a post-translational reporter cell line, referred to as MCF7-ERα-HiBiT, by fusing a small pro-luminescent nanoluciferase (NLuc) tag (HiBiT) to the C-terminus of endogenous ERα in MCF7 cells. The tag allows the luminescent detection and quantification of endogenous ERα protein by addition of the complementary NLuc enzyme fragment. This MCF7-ERα-HiBiT cell line was optimized for quantitative high-throughput screening (qHTS) to identify compounds that reduce ERα levels. In addition, the same cell line was optimized for a qHTS cellular thermal shift assay to identify compounds that bind and thermally stabilize ERα. Here, we interrogated the MCF7-ERα-HiBiT assay against the NCATS Pharmacological Collection (NPC) of 2,678 approved drugs and identified compounds that potently reduce and thermally stabilize ERα. Our novel post-translational reporter cell line provides a unique opportunity for profiling large pharmacological and toxicological compound libraries for their effect on ERα levels as well as for assessing direct compound binding to the receptor, thus facilitating mechanistic studies by which compounds exert their biological effects on ERα.

Effective treatments for COVID-19 are urgently needed. However, discovering single-agent therapies with activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been challenging. Combination therapies play an important role in antiviral therapies, due to their improved efficacy and reduced toxicity. Recent approaches have applied deep learning to identify synergistic drug combinations for diseases with vast preexisting datasets, but these are not applicable to new diseases with limited combination data, such as COVID-19. Given that drug synergy often occurs through inhibition of discrete biological targets, here we propose a neural network architecture that jointly learns drug-target interaction and drug-drug synergy. The model consists of two parts: a drug-target interaction module and a target-disease association module. This design enables the model to utilize drug-target interaction data and single-agent antiviral activity data, in addition to available drug-drug combination datasets, which may be small in nature. By incorporating additional biological information, our model performs significantly better in synergy prediction accuracy than previous methods with limited drug combination training data. We empirically validated our model predictions and discovered two drug combinations, remdesivir and reserpine as well as remdesivir and IQ-1S, which display strong antiviral SARS-CoV-2 synergy in vitro. Our approach, which was applied here to address the urgent threat of COVID-19, can be readily extended to other diseases for which a dearth of chemical-chemical combination data exists., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

The National Center for Advancing Translational Sciences (NCATS) has been actively generating SARS-CoV-2 high-throughput screening data and disseminates it through the OpenData Portal (https://opendata.ncats.nih.gov/covid19/). Here, we provide a hybrid approach that utilizes NCATS screening data from the SARS-CoV-2 cytopathic effect reduction assay to build predictive models, using both machine learning and pharmacophore-based modeling. Optimized models were used to perform two iterative rounds of virtual screening to predict small molecules active against SARS-CoV-2. Experimental testing with live virus provided 100 (∼16% of predicted hits) active compounds (efficacy > 30%, IC 50 ≤ 15 μM). Systematic clustering analysis of active compounds revealed three promising chemotypes which have not been previously identified as inhibitors of SARS-CoV-2 infection. Further investigation resulted in the identification of allosteric binders to host receptor angiotensin-converting enzyme 2; these compounds were then shown to inhibit the entry of pseudoparticles bearing spike protein of wild-type SARS-CoV-2, as well as South African B.1.351 and UK B.1.1.7 variants., Competing Interests: The authors declare no competing financial interest., (Not subject to U.S. Copyright. Published 2021 by American Chemical Society.)

COVID-19 has resulted in huge numbers of infections and deaths worldwide and brought the most severe disruptions to societies and economies since the Great Depression. Massive experimental and computational research effort to understand and characterize the disease and rapidly develop diagnostics, vaccines, and drugs has emerged in response to this devastating pandemic and more than 130 000 COVID-19-related research papers have been published in peer-reviewed journals or deposited in preprint servers. Much of the research effort has focused on the discovery of novel drug candidates or repurposing of existing drugs against COVID-19, and many such projects have been either exclusively computational or computer-aided experimental studies. Herein, we provide an expert overview of the key computational methods and their applications for the discovery of COVID-19 small-molecule therapeutics that have been reported in the research literature. We further outline that, after the first year the COVID-19 pandemic, it appears that drug repurposing has not produced rapid and global solutions. However, several known drugs have been used in the clinic to cure COVID-19 patients, and a few repurposed drugs continue to be considered in clinical trials, along with several novel clinical candidates. We posit that truly impactful computational tools must deliver actionable, experimentally testable hypotheses enabling the discovery of novel drugs and drug combinations, and that open science and rapid sharing of research results are critical to accelerate the development of novel, much needed therapeutics for COVID-19.

The effect of electron and proton acceptors on the photocyclization of diarylethenes has been studied. Without any additives, the deprotonation reaction is predominant, although other processes, including the sigmatropic shift, are not excluded. A deuterium exchange experiment has shown that a strong base (DABCO) facilitates the deprotonation reaction, thereby limiting the sigmatropic shift. In the presence of an oxidizing agent or additional sources of radicals (O 2 , I 2 , TEMPO), the processes of deprotonation and rearrangement (H-shift) are practically not observed, and the reaction proceeds along a radical pathway with the formation of phenanthrene or its heterocyclic analogue.

The aim of this paper is to investigate the response of a homogeneously aligned nematic nanosized hollow cavity (HANNHC) confined between two charged horizontal coaxial cylinders and subjected to both a radially applied electrostatic field E, arising from the surface charge density κ and the temperature gradient ∇T set between these cylinders. This was done within the framework of an extension of the classical Ericksen-Leslie theory, supplemented by thermomechanical correction of the shear stress and Rayleigh dissipation function, as well as taking into account the entropy balance equation. The physical mechanism responsible for the excitation of the hydrodynamic flow in the HANNHC is based on the interaction of the director and temperature gradients and the static electric field. Calculations show that under the influence of both the ∇T and E, a stationary flow u^{st} is excited in the HANNHC in the horizontal direction. It is shown that the electric force enforced by the flexoelectric polarization plays a crucial role in the excitation of u^{st} between these cylinders.

Several scenarios for the formation of hydrodynamic flows in microsized hybrid aligned nematic (HAN) channels, based on the appropriate nonlinear extension of the classical Ericksen-Leslie theory, supplemented by thermomechanical correction of the shear stress and Rayleigh dissipation function, as well as taking into account the entropy balance equation, are analyzed. Detailed numerical simulations were performed to elucidate the role of the heat flux q caused by laser radiation focused on the lower boundary of the equally warmed up the HAN channel containing a monolayer of azobenzene with the possibility of a trans-cis and cis-trans conformational changes in formation of the vortex flow v. It is shown that a thermally excited vortex flow is maintained with motion in a positive sense (clockwise) in the vicinity of the orientation defect at the lower boundary of the HAN channel caused by the trans-cis and cis-trans conformational changes. In the case of the same HAN channel, but without the azobenzene monolayer at the lower boundary, the heat flux q can also produce the vortical flow in the vicinity of the laser spot at the lower boundary, directed in a negative sense (counterclockwise). At that, the second vortex is characterized by a much slower speed than the vortical flow in the first case.

Activation-induced deaminase (AID) not only mutates DNA within the immunoglobulin loci to generate antibody diversity, but it also promotes development of B cell lymphomas. To tame this mutagen, we performed a quantitative high-throughput screen of over 90 000 compounds to see if AID activity could be mitigated. The enzymatic activity was assessed in biochemical assays to detect cytosine deamination and in cellular assays to measure class switch recombination. Three compounds showed promise via inhibition of switching in a transformed B cell line and in murine splenic B cells. These compounds have similar chemical structures, which suggests a shared mechanism of action. Importantly, the inhibitors blocked AID, but not a related cytosine DNA deaminase, APOBEC3B. We further determined that AID was continually expressed for several days after B cell activation to induce switching. This first report of small molecules that inhibit AID can be used to gain regulatory control over base editors., Competing Interests: The authors declare the following competing financial interest(s): R.S.H. is a co-founder, shareholder, and consultant of ApoGen Biotechnologies Inc., (© 2021 American Chemical Society.)

Endoplasmic reticulum (ER) dysregulation is associated with pathologies including neurodegenerative, muscular, and diabetic conditions. Depletion of ER calcium can lead to the loss of resident proteins in a process termed exodosis. To identify compounds that attenuate the redistribution of ER proteins under pathological conditions, we performed a quantitative high-throughput screen using the Gaussia luciferase (GLuc)-secreted ER calcium modulated protein (SERCaMP) assay, which monitors secretion of ER-resident proteins triggered by calcium depletion. We identify several clinically used drugs, including bromocriptine, and further characterize them using assays to measure effects on ER calcium, ER stress, and ER exodosis. Bromocriptine elicits protective effects in cell-based models of exodosis as well as in vivo models of stroke and diabetes. Bromocriptine analogs with reduced dopamine receptor activity retain similar efficacy in stabilizing the ER proteome, indicating a non-canonical mechanism of action. This study describes a strategic approach to identify small-molecule drugs capable of improving ER proteostasis in human disease conditions., Competing Interests: Declaration of interests F.U. is an inventor on US 10,441,574, B2 treatment for Wolfram syndrome and other ER stress disorders and US 9,891,231 B2 soluble MANF in pancreatic beta cell disorders. M.J.H., S.-M.Y., D.M., F.U., A.J., and B.K.H. are inventors on WO2018165171A1 treatment for Wolfram syndrome and other endoplasmic reticulum stress disorders., (Published by Elsevier Inc.)

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has prompted researchers to pivot their efforts to finding antiviral compounds and vaccines. In this study, we focused on the human host cell transmembrane protease serine 2 (TMPRSS2), which plays an important role in the viral life cycle by cleaving the spike protein to initiate membrane fusion. TMPRSS2 is an attractive target and has received attention for the development of drugs against SARS and Middle East respiratory syndrome. Starting with comparative structural modeling and a binding model analysis, we developed an efficient pharmacophore-based approach and applied a large-scale in silico database screening for small-molecule inhibitors against TMPRSS2. The hits were evaluated in the TMPRSS2 biochemical assay and the SARS-CoV-2 pseudotyped particle entry assay. A number of novel inhibitors were identified, providing starting points for the further development of drug candidates for the treatment of coronavirus disease 2019., Competing Interests: The authors declare no competing financial interest., (© 2021 American Chemical Society.)

Background: Humans are exposed to tens of thousands of chemical substances that need to be assessed for their potential toxicity. Acute systemic toxicity testing serves as the basis for regulatory hazard classification, labeling, and risk management. However, it is cost- and time-prohibitive to evaluate all new and existing chemicals using traditional rodent acute toxicity tests. In silico models built using existing data facilitate rapid acute toxicity predictions without using animals., Objectives: The U.S. Interagency Coordinating Committee on the Validation of Alternative Methods (ICCVAM) Acute Toxicity Workgroup organized an international collaboration to develop in silico models for predicting acute oral toxicity based on five different end points: Lethal Dose 50 ( LD 50 value, U.S. Environmental Protection Agency hazard (four) categories, Globally Harmonized System for Classification and Labeling hazard (five) categories, very toxic chemicals [ LD 50 ( LD 50 ≤ 50 mg / kg )], and nontoxic chemicals ( L D 50 > 2,000 mg / kg )., Methods: An acute oral toxicity data inventory for 11,992 chemicals was compiled, split into training and evaluation sets, and made available to 35 participating international research groups that submitted a total of 139 predictive models. Predictions that fell within the applicability domains of the submitted models were evaluated using external validation sets. These were then combined into consensus models to leverage strengths of individual approaches., Results: The resulting consensus predictions, which leverage the collective strengths of each individual model, form the Collaborative Acute Toxicity Modeling Suite (CATMoS). CATMoS demonstrated high performance in terms of accuracy and robustness when compared with in vivo results., Discussion: CATMoS is being evaluated by regulatory agencies for its utility and applicability as a potential replacement for in vivo rat acute oral toxicity studies. CATMoS predictions for more than 800,000 chemicals have been made available via the National Toxicology Program's Integrated Chemical Environment tools and data sets (ice.ntp.niehs.nih.gov). The models are also implemented in a free, standalone, open-source tool, OPERA, which allows predictions of new and untested chemicals to be made. https://doi.org/10.1289/EHP8495.

Computational methods to predict molecular properties regarding safety and toxicology represent alternative approaches to expedite drug development, screen environmental chemicals, and thus significantly reduce associated time and costs. There is a strong need and interest in the development of computational methods that yield reliable predictions of toxicity, and many approaches, including the recently introduced deep neural networks, have been leveraged towards this goal. Herein, we report on the collection, curation, and integration of data from the public data sets that were the source of the ChemIDplus database for systemic acute toxicity. These efforts generated the largest publicly available such data set comprising > 80,000 compounds measured against a total of 59 acute systemic toxicity end points. This data was used for developing multiple single- and multitask models utilizing random forest, deep neural networks, convolutional, and graph convolutional neural network approaches. For the first time, we also reported the consensus models based on different multitask approaches. To the best of our knowledge, prediction models for 36 of the 59 end points have never been published before. Furthermore, our results demonstrated a significantly better performance of the consensus model obtained from three multitask learning approaches that particularly predicted the 29 smaller tasks (less than 300 compounds) better than other models developed in the study. The curated data set and the developed models have been made publicly available at https://github.com/ncats/ld50-multitask, https://predictor.ncats.io/, and https://cactus.nci.nih.gov/download/acute-toxicity-db (data set only) to support regulatory and research applications.

Antiviral drug development for coronavirus disease 2019 (COVID-19) is occurring at an unprecedented pace, yet there are still limited therapeutic options for treating this disease. We hypothesized that combining drugs with independent mechanisms of action could result in synergy against SARS-CoV-2, thus generating better antiviral efficacy. Using in silico approaches, we prioritized 73 combinations of 32 drugs with potential activity against SARS-CoV-2 and then tested them in vitro. Sixteen synergistic and eight antagonistic combinations were identified; among 16 synergistic cases, combinations of the US Food and Drug Administration (FDA)-approved drug nitazoxanide with remdesivir, amodiaquine, or umifenovir were most notable, all exhibiting significant synergy against SARS-CoV-2 in a cell model. However, the combination of remdesivir and lysosomotropic drugs, such as hydroxychloroquine, demonstrated strong antagonism. Overall, these results highlight the utility of drug repurposing and preclinical testing of drug combinations for discovering potential therapies to treat COVID-19., (Copyright © 2020 The American Society of Gene and Cell Therapy. All rights reserved.)