TMV-CP based rational design and discovery of α-Amide phosphate derivatives as anti plant viral agents.

The tobacco mosaic virus coat protein (TMV-CP) is indispensable for the virus's replication, movement and transmission, as well as for the host plant's immune system to recognize it. It constitutes the outermost layer of the virus particle, and serves as an essential component of the virus structure. TMV-CP is essential for initiating and extending viral assembly, playing a crucial role in the self-assembly process of Tobacco Mosaic Virus (TMV). This research employed TMV-CP as a primary target for virtual screening, from which a library of 43,417 compounds was sourced and SH-05 was chosen as the lead compound. Consequently, a series of α-amide phosphate derivatives were designed and synthesized, exhibiting remarkable anti-TMV efficacy. The synthesized compounds were found to be beneficial in treating TMV, with compound 3g displaying a slightly better curative effect than Ningnanmycin (NNM) (EC50 = 304.54 µg/mL) at an EC50 of 291.9 µg/mL. Additionally, 3g exhibited comparable inactivation activity (EC50 = 63.2 µg/mL) to NNM (EC50 = 67.5 µg/mL) and similar protective activity (EC50 = 228.9 µg/mL) to NNM (EC50 = 219.7 µg/mL). Microscale thermal analysis revealed that the binding of 3g (Kd = 4.5 ± 1.9 µM) to TMV-CP showed the same level with NNM (Kd = 5.5 ± 2.6 µM). Results from transmission electron microscopy indicated that 3g could disrupt the structure of TMV virus particles. The toxicity prediction indicated that 3g was low toxicity. Molecular docking showed that 3g interacted with TMV-CP through hydrogen bond, attractive charge interaction and π-Cation interaction. This research provided a novel α-amide phosphate structure target TMV-CP, which may help the discovery of new anti-TMV agents in the future.

Structural and Functional Analyses of Inhibition of Human Dihydroorotate Dehydrogenase by Antiviral Furocoumavirin.

Natural products are important sources of seed compounds for drug discovery. However, it has become difficult in recent years to discover new compounds with valuable pharmacological activities. On the other hand, among the vast number of natural products that have been isolated so far, a considerable number of compounds with specific biological activities are thought to be overlooked in screening that uses biological activity as an index. Therefore, it is conceivable that such overlooked useful compounds may be found by screening compound libraries that have been amassed previously through specific assays. Previously, NPD723, a member of the Natural Products Depository library comprised of a mixture of natural and non-natural products developed at RIKEN, and its metabolite H-006 were found to inhibit growth of various cancer cells at low nanomolar half-maximal inhibitory concentration. Subsequent analysis revealed that H-006 strongly inhibited human dihydroorotate dehydrogenase (DHODH), the rate-limiting enzyme in the de novo pyrimidine biosynthetic pathway. Here, we elucidated the crystal structure of the DHODH-flavin mononucleotide-orotic acid-H-006 complex at 1.7 Å resolution to determine that furocoumavirin, the S-enantiomer of H-006, was the actual inhibitor. The overall mode of interaction of furocoumavirin with the inhibitor binding pocket was similar to that described for previously reported tight-binding inhibitors. However, the structural information together with kinetic characterizations of site-specific mutants identified key unique features that are considered to contribute to the sub-nanomolar inhibition of DHODH by furocoumavirin. Our finding identified new chemical features that could improve the design of human DHODH inhibitors.

Bioactive Lyocell Fibers with Inherent Antibacterial, Antiviral and Antifungal Properties.

Functional Lyocell fibers gain interest in garments and technical textiles, especially when equipped with inherently bioactive features. In this study, Lyocell fibers are modified with an ion exchange resin and subsequently loaded with copper (Cu) ions. The modified Lyocell process enables high amounts of the resin additive (>10%) through intensive dispersion and subsequently, high uptake of 2.7% Cu throughout the whole cross-section of the fiber. Fixation by Na2CO3 increases the washing and dyeing resistance considerably. Cu content after dyeing compared to the original fiber value amounts to approx. 65% for reactive, 75% for direct, and 77% for HT dyeing, respectively. Even after 50 household washes, a recovery of 43% for reactive, 47% for direct and 26% for HT dyeing is proved. XRD measurements reveal ionic bonding of Cu fixation inside the cellulose/ion exchange resin composite. A combination of the fixation process with a change in Cu valence state by glucose/NaOH leads to the formation of Cu2O crystallites, which is proved by XRD. Cu fiber shows a strong antibacterial effect against Staphylococcus aureus and Klebsiella pneumonia bacteria, even after 50 household washing cycles of both >5 log CFU. In nonwoven blends with a share of only 6% Cu fiber, a strong antimicrobial (CFU > log 5) and full antiviral effectiveness (>log 4) was received even after 50 washing cycles. Time-dependent measurements already show strong antiviral behavior after 30 s. Further, the fibers show an increased die off of the fungal isolate Candida auris with CFU log 4.4, and nonwovens made from 6% Cu fiber share a CFU log of 1.7. Findings of the study predestines the fiber for advanced textile processing and applications in areas with high germ loads.

Drug repurposing: identification of SARS-CoV-2 potential inhibitors by virtual screening and pharmacokinetics strategies.

INTRODUCTION: The coronavirus disease 2019 (COVID-19) pandemic caused global health, economic, and population loss. Variants of the coronavirus contributed to the severity of the disease and persistent rise in infections. This study aimed to identify potential drug candidates from fifteen approved antiviral drugs against SARS-CoV-2 (6LU7), SARS-CoV (5B6O), and SARS-CoV-2 spike protein (6M0J) using virtual screening and pharmacokinetics to gain insights into COVID-19 therapeutics. METHODOLOGY: We employed drug repurposing approach to analyze binding performance of fifteen clinically approved antiviral drugs against the main protease of SARS-CoV-2 (6LU7), SARS-CoV (5B6O), and SARS-CoV-2 spike proteins bound to ACE-2 receptor (6M0J), to provide an insight into the therapeutics of COVID-19. AutoDock Vina was used for docking studies. The binding affinities were calculated, and 2-3D structures of protein-ligand interactions were drawn. RESULTS: Rutin, hesperidin, and nelfinavir are clinically approved antiviral drugs with high binding affinity to proteins 6LU7, 5B6O, and 6M0J. These ligands have excellent pharmacokinetics, ensuring efficient absorption, metabolism, excretion, and digestibility. Hesperidin showed the most potent interaction with spike protein 6M0J, forming four H-bonds. Nelfinavir had a high human intestinal absorption (HIA) score of 0.93, indicating maximum absorption in the body and promising interactions with 6LU7. CONCLUSIONS: Our results indicated that rutin, hesperidin, and nelfinavir had the highest binding results against the proposed drug targets. The computational approach effectively identified SARS-CoV-2 inhibitors. COVID-19 is still a recurrent threat globally and predictive analysis using natural compounds might serve as a starting point for new drug development against SARS-CoV-2 and related viruses.

An Overview of SARS-CoV-2 Potential Targets, Inhibitors, and Computational Insights to Enrich the Promising Treatment Strategies.

The rapid spread of the SARS-CoV-2 virus has emphasized the urgent need for effective therapies to combat COVID-19. Investigating the potential targets, inhibitors, and in silico approaches pertinent to COVID-19 are of utmost need to develop novel therapeutic agents and reprofiling of existing FDA-approved drugs. This article reviews the viral enzymes and their counter receptors involved in the entry of SARS-CoV-2 into host cells, replication of genomic RNA, and controlling the host cell physiology. In addition, the study provides an overview of the computational techniques such as docking simulations, molecular dynamics, QSAR modeling, and homology modeling that have been used to find the FDA-approved drugs and other inhibitors against SARS-CoV-2. Furthermore, a comprehensive overview of virus-based and host-based druggable targets from a structural point of view, together with the reported therapeutic compounds against SARS-CoV-2 have also been presented. The current study offers future perspectives for research in the field of network pharmacology investigating the large unexplored molecular libraries. Overall, the present in-depth review aims to expedite the process of identifying and repurposing drugs for researchers involved in the field of COVID-19 drug discovery.

Exploring the Antiviral Potential of Natural Compounds against Influenza: A Combined Computational and Experimental Approach.

The influenza A virus nonstructural protein 1 (NS1), which is crucial for viral replication and immune evasion, has been identified as a significant drug target with substantial potential to contribute to the fight against influenza. The emergence of drug-resistant influenza A virus strains highlights the urgent need for novel therapeutics. This study proposes a combined theoretical criterion for the virtual screening of molecular libraries to identify candidate NS1 inhibitors. By applying the criterion to the ZINC Natural Product database, followed by ligand-based virtual screening and molecular docking, we proposed the most promising candidate as a potential NS1 inhibitor. Subsequently, the selected natural compound was experimentally evaluated, revealing measurable virus replication inhibition activity in cell culture. This approach offers a promising avenue for developing novel anti-influenza agents targeting the NS1 protein.

Exploring Binding Pockets in the Conformational States of the SARS-CoV-2 Spike Trimers for the Screening of Allosteric Inhibitors Using Molecular Simulations and Ensemble-Based Ligand Docking.

Understanding mechanisms of allosteric regulation remains elusive for the SARS-CoV-2 spike protein, despite the increasing interest and effort in discovering allosteric inhibitors of the viral activity and interactions with the host receptor ACE2. The challenges of discovering allosteric modulators of the SARS-CoV-2 spike proteins are associated with the diversity of cryptic allosteric sites and complex molecular mechanisms that can be employed by allosteric ligands, including the alteration of the conformational equilibrium of spike protein and preferential stabilization of specific functional states. In the current study, we combine conformational dynamics analysis of distinct forms of the full-length spike protein trimers and machine-learning-based binding pocket detection with the ensemble-based ligand docking and binding free energy analysis to characterize the potential allosteric binding sites and determine structural and energetic determinants of allosteric inhibition for a series of experimentally validated allosteric molecules. The results demonstrate a good agreement between computational and experimental binding affinities, providing support to the predicted binding modes and suggesting key interactions formed by the allosteric ligands to elicit the experimentally observed inhibition. We establish structural and energetic determinants of allosteric binding for the experimentally known allosteric molecules, indicating a potential mechanism of allosteric modulation by targeting the hinges of the inter-protomer movements and blocking conformational changes between the closed and open spike trimer forms. The results of this study demonstrate that combining ensemble-based ligand docking with conformational states of spike protein and rigorous binding energy analysis enables robust characterization of the ligand binding modes, the identification of allosteric binding hotspots, and the prediction of binding affinities for validated allosteric modulators, which is consistent with the experimental data. This study suggested that the conformational adaptability of the protein allosteric sites and the diversity of ligand bound conformations are both in play to enable efficient targeting of allosteric binding sites and interfere with the conformational changes.

In Vitro Effect of 9,9'-Norharmane Dimer against Herpes Simplex Viruses.

Herpes simplex virus (HSV) infections are highly widespread among humans, producing symptoms ranging from ulcerative lesions to severe diseases such as blindness and life-threatening encephalitis. At present, there are no vaccines available, and some existing antiviral treatments can be ineffective or lead to adverse effects. As a result, there is a need for new anti-HSV drugs. In this report, the in vitro anti-HSV effect of 9,9'-norharmane dimer (nHo-dimer), which belongs to the ß-carboline (ßC) alkaloid family, was evaluated. The dimer exhibited no virucidal properties and did not impede either the attachment or penetration steps of viral particles. The antiviral effect was only exerted under the constant presence of the dimer in the incubation media, and the mechanism of action was found to involve later events of virus infection. Analysis of fluorescence lifetime imaging data showed that the nHo-dimer internalized well into the cells when present in the extracellular incubation medium, with a preferential accumulation into perinuclear organelles including mitochondria. After washing the host cells with fresh medium free of nHo-dimer, the signal decreased, suggesting the partial release of the compound from the cells. This agrees with the observation that the antiviral effect is solely manifested when the alkaloid is consistently present in the incubation media.

Nanoformulation of the Broad-Spectrum Hydrophobic Antiviral Vacuolar ATPase Inhibitor Diphyllin in Human Recombinant H-ferritin.

Background: As highlighted by recent pandemic outbreaks, antiviral drugs are crucial resources in the global battle against viral diseases. Unfortunately, most antiviral drugs are characterized by a plethora of side effects and low efficiency/poor bioavailability owing to their insolubility. This also applies to the arylnaphthalide lignin family member, diphyllin (Diph). Diph acts as a vacuolar ATPase inhibitor and has been previously identified as a promising candidate with broad-spectrum antiviral activity. However, its physicochemical properties preclude its efficient administration in vivo, complicating preclinical testing. Methods: We produced human recombinant H- ferritin (HsaFtH) and used it as a delivery vehicle for Diph encapsulation through pH-mediated reversible reassembly of HsaFtH. Diph nanoformulation was subsequently thoroughly characterized and tested for its non-target cytotoxicity and antiviral efficiency using a panel of pathogenic viral strain. Results: We revealed that loading into HsaFtH decreased the undesired cytotoxicity of Diph in mammalian host cells. We also confirmed that encapsulated Diph exhibited slightly lower antiviral activity than free Diph, which may be due to the differential uptake mechanism and kinetics of free Diph and Diph@HsaFtH. Furthermore, we confirmed that the antiviral effect was mediated solely by Diph with no contribution from HsaFtH. Conclusion: It was confirmed that HsaFtH is a suitable vehicle that allows easy loading of Diph and production of highly homogeneous nanoparticles dispersion with promising broad-spectrum antiviral activity.

Anti-respiratory syncytial virus and anti-herpes simplex virus activity of chemically engineered sulfated fucans from Cystoseira indica.

Seaweed polysaccharides, particularly sulfated ones, exhibited potent antiviral activity against a wide variety of enveloped viruses, such as herpes simplex virus and respiratory viruses. Different mechanisms of action were suggested, which may range from preventing infection to intracellular antiviral activity, at different stages of the viral cycle. Herein, we generated two chemically engineered sulfated fucans (C303 and C304) from Cystoseira indica by an amalgamated extraction-sulfation procedure using chlorosulfonic acid-pyridine/N,N-dimethylformamide and sulfur trioxide-pyridine/N,N-dimethylformamide reagents, respectively. These compounds exhibited activity against HSV-1 and RSV with 50 % inhibitory concentration values in the range of 0.75-2.5 µg/mL and low cytotoxicity at concentrations up to 500 µg/mL. The antiviral activities of chemically sulfated fucans (C303 and C304) were higher than the water (C301) and CaCl2 extracted (C302) polysaccharides. Compound C303 had a (1,3)-linked fucan backbone and was branched. Sulfates were present at positions C-2, C-4, and C-2,4 of Fucp, and C-6 of Galp residues of this polymer. Compound C304 had a comparable structure but with more sulfates at C-4 of Fucp residue. Both C303 and C304 were potent antiviral candidates, acting in a dose-dependent manner on the adsorption and other intracellular stages of HSV-1 and RSV replication, in vitro.

Seaweed-derived fucoidans and rhamnan sulfates serve as potent anti-SARS-CoV-2 agents with potential for prophylaxis.

Seaweeds represent a rich source of sulfated polysaccharides with similarity to heparan sulfate, a facilitator of myriad virus host cell attachment. For this reason, attention has been drawn to their antiviral activity, including the potential for anti-SARS-CoV-2 activity. We have identified and structurally characterized several fucoidan extracts, including those from different species of brown macroalga, and a rhamnan sulfate from a green macroalga species. A high molecular weight fucoidan extracted from Saccharina japonica (FSjRPI-27), and a rhamnan sulfate extracted from Monostroma nitidum (RSMn), showed potent competitive inhibition of spike glycoprotein receptor binding to a heparin-coated SPR chip. This inhibition was also observed in cell-based assays using hACE2 HEK-293 T cells infected by pseudotyped SARS-CoV-2 virus with IC50 values <1 µg/mL. Effectiveness was demonstrated in vivo using hACE2-transgenic mice. Intranasal administration of FSjRPI-27 showed protection when dosed 6 h prior to and at infection, and then every 2 days post-infection, with 100 % survival and no toxicity at 104 plaque-forming units per mouse vs. buffer control. At 5-fold higher virus dose, FSjRPI-27 reduced mortality and yielded reduced viral titers in bronchioalveolar fluid and lung homogenates vs. buffer control. These findings suggest the potential application of seaweed-based sulfated polysaccharides as promising anti-SARS-CoV-2 prophylactics.

Skin- and Eco-Friendly Hand Sanitizer: A Novel Composition of Natural Extracts to Prevent the Spread of Respiratory Viruses.

Respiratory diseases caused by viruses are a serious global health threat. Although the use of hand sanitizers containing alcohol and synthetic antiseptic agents is recognized as an effective, simple, and low-cost measure to combat viral transmission, they can harm human health and the environment. Thus, this work aimed to study the efficacy of combining Camellia sinensis and Chamomilla recutita extracts in a skin- and eco-friendly leave-on hand sanitizer to prevent the spread of respiratory viruses. An oil-in-water emulsion containing C. recutita oily extract (5.0%), C. recutita glycolic extract (0.2%) and C. sinensis glycolic extract (5.0%) showed virucidal activity against HAdV-2 (respiratory virus) and two surrogate viruses of SARS-CoV-2 (HSV-1 and MVH-3), showing great potential to prevent the spread of respiratory viruses. These natural extracts combined are also promising to combat a broad spectrum of other viruses, in the form of antiseptic mouthwashes or throat sprays, surface disinfectants, and veterinary products, among others. Complementally, the developed hand sanitizer demonstrated efficacy against bacteria and fungus.

Differential Bioactivation Profiles of Different GS-441524 Prodrugs in Cell and Mouse Models: ProTide Prodrugs with High Cell Permeability and Susceptibility to Cathepsin A Are More Efficient in Delivering Antiviral Active Metabolites to the Lung.

We assessed factors that determine the tissue-specific bioactivation of ProTide prodrugs by comparing the disposition and activation of remdesivir (RDV), its methylpropyl and isopropyl ester analogues (MeRDV and IsoRDV, respectively), the oral prodrug GS-621763, and the parent nucleotide GS-441524 (Nuc). RDV and MeRDV yielded more active metabolite remdesivir-triphosphate (RDV-TP) than IsoRDV, GS-621763, and Nuc in human lung cell models due to superior cell permeability and higher susceptivity to cathepsin A. Intravenous administration to mice showed that RDV and MeRDV delivered significantly more RDV-TP to the lung than other compounds. Nevertheless, all four ester prodrugs exhibited very low oral bioavailability (<2%), with Nuc being the predominant metabolite in blood. In conclusion, ProTides prodrugs, such as RDV and MeRDV, are more efficient in delivering active metabolites to the lung than Nuc, driven by high cell permeability and susceptivity to cathepsin A. Optimizing ProTides' ester structures is an effective strategy for enhancing prodrug activation in the lung.

An Elastin-like Polypeptide-fusion peptide targeting capsid-tegument interface as an antiviral against cytomegalovirus infection.

The tegument protein pp150 of Human Cytomegalovirus (HCMV) is known to be essential for the final stages of virus maturation and mediates its functions by interacting with capsid proteins. Our laboratory has previously identified the critical regions in pp150 important for pp150-capsid interactions and designed peptides similar in sequence to these regions, with a goal to competitively inhibit capsid maturation. Treatment with a specific peptide (PepCR2 or P10) targeted to pp150 conserved region 2 led to a significant reduction in murine CMV (MCMV) growth in cell culture, paving the way for in vivo testing in a mouse model of CMV infection. However, the general pharmacokinetic parameters of peptides, including rapid degradation and limited tissue and cell membrane permeability, pose a challenge to their successful use in vivo. Therefore, we designed a biopolymer-stabilized elastin-like polypeptide (ELP) fusion construct (ELP-P10) to enhance the bioavailability of P10. Antiviral efficacy and cytotoxic effects of ELP-P10 were studied in cell culture, and pharmacokinetics, biodistribution, and antiviral efficacy were studied in a mouse model of CMV infection. ELP-P10 maintained significant antiviral activity in cell culture, and this conjugation significantly enhanced P10 bioavailability in mouse tissues. The fluorescently labeled ELP-P10 accumulated to higher levels in mouse liver and kidneys as compared to the unconjugated P10. Moreover, viral titers from vital organs of MCMV-infected mice indicated a significant reduction of virus load upon ELP-P10 treatment. Therefore, ELP-P10 has the potential to be developed into an effective antiviral against CMV infection.

A Comprehensive Update of Anti-COVID-19 Activity of Heterocyclic Compounds.

The Coronavirus disease 2019 (COVID-19) pandemic is one of the most considerable health problems across the world. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the major causative agent of COVID-19. The severe symptoms of this deadly disease include shortness of breath, fever, cough, loss of smell, and a broad spectrum of other health issues such as diarrhea, pneumonia, bronchitis, septic shock, and multiple organ failure. Currently, there are no medications available for coronavirus patients, except symptom-relieving drugs. Therefore, SARS-CoV-2 requires the development of effective drugs and specific treatments. Heterocycles are important constituents of more than 85% of the physiologically active pharmaceutical drugs on the market now. Several FDA-approved drugs have been reported including molnupiravir, remdesivir, ritonavir, oseltamivir, favipiravir, chloroquine, and hydroxychloroquine for the cure of COVID-19. In this study, we discuss potent anti-SARS-CoV-2 heterocyclic compounds that have been synthesized over the past few years. These compounds included; indole, piperidine, pyrazine, pyrimidine, pyrrole, piperazine, quinazoline, oxazole, quinoline, isoxazole, thiazole, quinoxaline, pyrazole, azafluorene, imidazole, thiadiazole, triazole, coumarin, chromene, and benzodioxole. Both in vitro and in silico studies were performed to determine the potential of these heterocyclic compounds in the fight against various SARS-CoV-2 proteins.

Microwave-assisted synthesis of highly sulfated mannuronate glycans as potential inhibitors against SARS-CoV-2.

Algae-based marine carbohydrate drugs are typically decorated with negative ion groups such as carboxylate and sulfate groups. However, the precise synthesis of highly sulfated alginates is challenging, thus impeding their structure-activity relationship studies. Herein we achieve a microwave-assisted synthesis of a range of highly sulfated mannuronate glycans with up to 17 sulfation sites by overcoming the incomplete sulfation due to the electrostatic repulsion of crowded polyanionic groups. Although the partially sulfated tetrasaccharide had the highest affinity for the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant, the fully sulfated octasaccharide showed the most potent interference with the binding of the RBD to angiotensin-converting enzyme 2 (ACE2) and Vero E6 cells, indicating that the sulfated oligosaccharides might inhibit the RBD binding to ACE2 in a length-dependent manner.

Repairing Host Damage Caused by Tobacco Mosaic Virus Stress: Design, Synthesis, and Mechanism Study of Novel Oxadiazole and Arylhydrazone Derivatives.

Tobacco mosaic virus (TMV), as one of the most traditional and extensive biological stresses, poses a serious threat to plant growth and development. In this work, a series of 1-phenyl/tertbutyl-5-amino-4-pyrazole oxadiazole and arylhydrazone derivatives was synthesized. Bioassay evaluation demonstrated that the title compounds (P1-P18) without a "thioether bond" lost their anti-TMV activity, while some of the ring-opening arylhydrazone compounds exhibited superior in vivo activity against TMV in tobacco. The EC50 value of title compound T8 for curative activity was 139 µg/mL, similar to that of ningnanmycin (NNM) (EC50 = 152 µg/mL). Safety analysis revealed that compound T8 had no adverse effects on plant growth or seed germination at a concentration of 250 µg/mL. Morphological observation revealed that compound T8 could restore the leaf tissue of a TMV-stressed host and the leaf stomatal aperture to normal. A mechanism study further revealed that compound T8 not only restored the photosynthetic and growth ability of the damaged host to normal levels but also enhanced catalase (CAT) activity and reduced the content of malondialdehyde (MDA) and hydrogen peroxide (H2O2) in the damaged host, thereby reducing the oxidation damage to the host. TMV-green fluorescent protein (GFP) experiments further demonstrated that compound T8 not only slowed the transmission speed of TMV in the host but also inhibited its reproduction. All of the experimental results demonstrated that compound T8 could reduce the oxidative damage caused by TMV stress and regulate the photosynthetic ability of the host, achieving the ability to repair damage, to make the plant grow normally.

A cell based assay using virus-like particles to screen AM type mimics for SARS-CoV-2 neutralisation.

A number of small molecule and protein therapeutic candidates have been developed in the last four years against SARS-CoV-2 spike. However, there are hardly a few molecules that have advanced through the subsequent discovery steps to eventually work as a therapeutic agent. This is majorly because of the hurdles in determining the affinity of potential therapeutics with live SARS-CoV-2 virus. Furthermore, affinity determined for the receptor binding domain (RBD) of the SARS-CoV-2 spike protein, at times, fails to mimic physiological conditions of the host-virus interaction. To bridge this gap between in vitro and in vivo methods of therapeutic agent screening, we report an improved screening protocol for therapeutic candidates using SARS-CoV-2 virus like particles (VLPs). To minimise the interference from the bulkier reporters like GPF in the affinity studies, a smaller hemagglutinin (HA) tag has been fused to one of the proteins of VLP. This HA tag serves as readout, when probed with fluorescent anti-HA antibodies. Outcome of this study sheds light on the lesser known virus neutralisation capabilities of AM type miniprotein mimics. Further, to assess the stability of SARS-CoV-2 spike - miniprotein complex, we have performed molecular dynamic simulations on the membrane embedded protein complex. Simulation results reveal extremely stable intermolecular interactions between RBD and one of the AM type miniproteins, AM1. Furthermore, we discovered a robust network of intramolecular interactions that help stabilise AM1. Findings from our in vitro and in silico experiments concurrently highlight advantages and capabilities of mimic based miniprotein therapeutics.

Allosteric inhibition of dengue virus RNA-dependent RNA polymerase by Litsea cubeba phytochemicals: a computational study.

RNA-dependent RNA polymerase (RdRp) is considered a potential drug target for dengue virus (DENV) inhibition and has attracted attention in antiviral drug discovery. Here, we screened 121 natural compounds from Litsea cubeba against DENV RdRp using various approaches of computer-based drug discovery. Notably, we identified four potential compounds (Ushinsunine, Cassameridine, (+)-Epiexcelsin, (-)-Phanostenine) with good binding scores and allosteric interactions with the target protein. Moreover, molecular dynamics simulation studies were done to check the conformational stability of the complexes under given conditions. Additionally, we performed post-simulation analysis to find the stability of potential drugs in the target protein. The findings suggest Litsea cubeba-derived phytomolecules as a therapeutic solution to control DENV infection.Communicated by Ramaswamy H. Sarma.

Phytoconstituents of Artemisia Annua as potential inhibitors of SARS CoV2 main protease: an in silico study.

BACKGROUND: In November 2019, the world faced a pandemic called SARS-CoV-2, which became a major threat to humans and continues to be. To overcome this, many plants were explored to find a cure. METHODS: Therefore, this research was planned to screen out the active constituents from Artemisia annua that can work against the viral main protease Mpro as this non-structural protein is responsible for the cleavage of replicating enzymes of the virus. Twenty-five biocompounds belonging to different classes namely alpha-pinene, beta-pinene, carvone, myrtenol, quinic acid, caffeic acid, quercetin, rutin, apigenin, chrysoplenetin, arteannunin b, artemisinin, scopoletin, scoparone, artemisinic acid, deoxyartemisnin, artemetin, casticin, sitogluside, beta-sitosterol, dihydroartemisinin, scopolin, artemether, artemotil, artesunate were selected. Virtual screening of these ligands was carried out against drug target Mpro by CB dock. RESULTS: Quercetin, rutin, casticin, chrysoplenetin, apigenin, artemetin, artesunate, sopolin and sito-gluside were found as hit compounds. Further, ADMET screening was conducted which represented Chrysoplenetin as a lead compound. Azithromycin was used as a standard drug. The interactions were studied by PyMol and visualized in LigPlot. Furthermore, the RMSD graph shows fluctuations at various points at the start of simulation in Top1 (Azithromycin) complex system due to structural changes in the helix-coil-helix and beta-turn-beta changes at specific points resulting in increased RMSD with a time frame of 50 ns. But this change remains stable after the extension of simulation time intervals till 100 ns. On other side, the Top2 complex system remains highly stable throughout the time scale. No such structural dynamics were observed bu the ligand attached to the active site residues binds strongly. CONCLUSION: This study facilitates researchers to develop and discover more effective and specific therapeutic agents against SARS-CoV-2 and other viral infections. Finally, chrysoplenetin was identified as a more potent drug candidate to act against the viral main protease, which in the future can be helpful.