Thapsigargin and Tunicamycin Block SARS-CoV-2 Entry into Host Cells via Differential Modulation of Unfolded Protein Response (UPR), AKT Signaling, and Apoptosis.

BACKGROUND: SARS-Co-V2 infection can induce ER stress-associated activation of unfolded protein response (UPR) in host cells, which may contribute to the pathogenesis of COVID-19. To understand the complex interplay between SARS-Co-V2 infection and UPR signaling, we examined the effects of acute pre-existing ER stress on SARS-Co-V2 infectivity. METHODS: Huh-7 cells were treated with Tunicamycin (TUN) and Thapsigargin (THA) prior to SARS-CoV-2pp transduction (48 h p.i.) to induce ER stress. Pseudo-typed particles (SARS-CoV-2pp) entry into host cells was measured by Bright GloTM luciferase assay. Cell viability was assessed by cell titer Glo® luminescent assay. The mRNA and protein expression was evaluated by RT-qPCR and Western Blot. RESULTS: TUN (5 µg/mL) and THA (1 µM) efficiently inhibited the entry of SARS-CoV-2pp into host cells without any cytotoxic effect. TUN and THA's attenuation of virus entry was associated with differential modulation of ACE2 expression. Both TUN and THA significantly reduced the expression of stress-inducible ER chaperone GRP78/BiP in transduced cells. In contrast, the IRE1-XBP1s and PERK-eIF2α-ATF4-CHOP signaling pathways were downregulated with THA treatment, but not TUN in transduced cells. Insulin-mediated glucose uptake and phosphorylation of Ser307 IRS-1 and downstream p-AKT were enhanced with THA in transduced cells. Furthermore, TUN and THA differentially affected lipid metabolism and apoptotic signaling pathways. CONCLUSIONS: These findings suggest that short-term pre-existing ER stress prior to virus infection induces a specific UPR response in host cells capable of counteracting stress-inducible elements signaling, thereby depriving SARS-Co-V2 of essential components for entry and replication. Pharmacological manipulation of ER stress in host cells might provide new therapeutic strategies to alleviate SARS-CoV-2 infection.

Predicting drug-Protein interaction with deep learning framework for molecular graphs and sequences: Potential candidates against SAR-CoV-2.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the COVID-19 disease, which represents a new life-threatening disaster. Regarding viral infection, many therapeutics have been investigated to alleviate the epidemiology such as vaccines and receptor decoys. However, the continuous mutating coronavirus, especially the variants of Delta and Omicron, are tended to invalidate the therapeutic biological product. Thus, it is necessary to develop molecular entities as broad-spectrum antiviral drugs. Coronavirus replication is controlled by the viral 3-chymotrypsin-like cysteine protease (3CLpro) enzyme, which is required for the virus's life cycle. In the cases of severe acute respiratory syndrome coronavirus (SARS-CoV) and middle east respiratory syndrome coronavirus (MERS-CoV), 3CLpro has been shown to be a promising therapeutic development target. Here we proposed an attention-based deep learning framework for molecular graphs and sequences, training from the BindingDB 3CLpro dataset (114,555 compounds). After construction of such model, we conducted large-scale screening the in vivo/vitro dataset (276,003 compounds) from Zinc Database and visualize the candidate compounds with attention score. geometric-based affinity prediction was employed for validation. Finally, we established a 3CLpro-specific deep learning framework, namely GraphDPI-3CL (AUROC: 0.958) achieved superior performance beyond the existing state of the art model and discovered 10 molecules with a high binding affinity of 3CLpro and superior binding mode.

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.

Antiviral, anti-inflammatory and antioxidant effects of curcumin and curcuminoids in SH-SY5Y cells infected by SARS-CoV-2.

COVID-19, caused by SARS-CoV-2, affects neuronal cells, causing several symptoms such as memory loss, anosmia and brain inflammation. Curcuminoids (Me08 e Me23) and curcumin (CUR) are derived from Curcuma Longa extract (EXT). Many therapeutic actions have been linked to these compounds, including antiviral action. Given the severe implications of COVID-19, especially within the central nervous system, our study aims to shed light on the therapeutic potential of curcuminoids against SARS-CoV-2 infection, particularly in neuronal cells. Here, we investigated the effects of CUR, EXT, Me08 and Me23 in human neuroblastoma SH-SY5Y. We observed that Me23 significantly decreased the expression of plasma membrane-associated transmembrane protease serine 2 (TMPRSS2) and TMPRSS11D, consequently mitigating the elevated ROS levels induced by SARS-CoV-2. Furthermore, Me23 exhibited antioxidative properties by increasing NRF2 gene expression and restoring NQO1 activity following SARS-CoV-2 infection. Both Me08 and Me23 effectively reduced SARS-CoV-2 replication in SH-SY5Y cells overexpressing ACE2 (SH-ACE2). Additionally, all of these compounds demonstrated the ability to decrease proinflammatory cytokines such as IL-6, TNF-α, and IL-17, while Me08 specifically reduced INF-γ levels. Our findings suggest that curcuminoid Me23 could serve as a potential agent for mitigating the impact of COVID-19, particularly within the context of central nervous system involvement.

A Multi-Faceted Binding Assessment of Aptamers Targeting the SARS-CoV-2 Spike Protein.

The COVID-19 pandemic has underscored the critical need for the advancement of diagnostic and therapeutic platforms. These platforms rely on the rapid development of molecular binders that should facilitate surveillance and swift intervention against viral infections. In this study, we have evaluated by three independent research groups the binding characteristics of various published RNA and DNA aptamers targeting the spike protein of the SARS-CoV-2 virus. For this comparative analysis, we have employed different techniques such as biolayer interferometry (BLI), enzyme-linked oligonucleotide assay (ELONA), and flow cytometry. Our data show discrepancies in the reported specificity and affinity among several of the published aptamers and underline the importance of standardized methods, the impact of biophysical techniques, and the controls used for aptamer characterization. We expect our results to contribute to the selection and application of suitable aptamers for the detection of SARS-CoV-2.

Collection and transportation of SARS-CoV-2 and influenza A virus diagnostic samples: Optimizing the usage of guanidine-based chaotropic salts for enhanced biosafety and viral genome preservation.

Many virus lysis/transport buffers used in molecular diagnostics, including the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA, contain guanidine-based chaotropic salts, primarily guanidine hydrochloride (GuHCl) or guanidine isothiocyanate (GITC). Although the virucidal effects of GuHCl and GITC alone against some enveloped viruses have been established, standardized data on their optimum virucidal concentrations against SARS-CoV-2 and effects on viral RNA stability are scarce. Thus, we aimed to determine the optimum virucidal concentrations of GuHCl and GITC against SARS-CoV-2 compared to influenza A virus (IAV), another enveloped respiratory virus. We also evaluated the effectiveness of viral RNA stabilization at the determined optimum virucidal concentrations under high-temperature conditions (35°C) using virus-specific real-time reverse transcription polymerase chain reaction. Both viruses were potently inactivated by 1.0 M GITC and 2.5 M GuHCl, but the GuHCl concentration for efficient SARS-CoV-2 inactivation was slightly higher than that for IAV inactivation. GITC showed better viral RNA stability than GuHCl at the optimum virucidal concentrations. An increased concentration of GuHCl or GITC increased viral RNA degradation at 35°C. Our findings highlight the need to standardize GuHCl and GITC concentrations in virus lysis/transport buffers and the potential application of these guanidine-based salts alone as virus inactivation solutions in SARS-CoV-2 and IAV molecular diagnostics.

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.

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.

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.

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.

Changing patterns of bacterial profile and antimicrobial resistance in high-risk patients during the COVID-19 pandemic at a tertiary oncology hospital.

The widespread evolution of phenotypic resistance in clinical isolates over the years, coupled with the COVID-19 pandemic onset, has exacerbated the global challenge of antimicrobial resistance. This study aimed to explore changes in bacterial infection patterns and antimicrobial resistance during the COVID-19 pandemic. This study involved the periods before and during COVID-19: the pre-pandemic and pandemic eras. The surveillance results of bacterial isolates causing infections in cancer patients at an Egyptian tertiary oncology hospital were retrieved. The Vitek®2 or Phoenix systems were utilized for species identification and susceptibility testing. Statistical analyses were performed comparing microbiological trends before and during the pandemic. Out of 2856 bacterial isolates, Gram-negative bacteria (GNB) predominated (69.7%), and Gram-positive bacteria (GPB) comprised 30.3% of isolates. No significant change was found in GNB prevalence during the pandemic (P = 0.159). Elevated rates of Klebsiella and Pseudomonas species were demonstrated during the pandemic, as was a decrease in E. coli and Acinetobacter species (P < 0.001, 0.018, < 0.001, and 0.046, respectively) in hematological patients. In surgical patients, Enterobacteriaceae significantly increased (P = 0.012), while non-fermenters significantly decreased (P = 0.007). GPB species from either hematological or surgical wards exhibited no notable changes during the pandemic. GNB resistance increased in hematological patients to carbapenems, amikacin, and tigecycline and decreased in surgical patients to amikacin and cefoxitin (P < 0.001, 0.010, < 0.001, < 0.001, and 0.016, respectively). The study highlights notable shifts in the microbial landscape during the COVID-19 pandemic, particularly in the prevalence and resistance patterns of GNB in hematological and surgical wards.

Synthesis and in silico inhibitory action studies of azo-anchored imidazo[4,5-b]indole scaffolds against the COVID-19 main protease (Mpro).

The present work elicits a novel approach to combating COVID-19 by synthesizing a series of azo-anchored 3,4-dihydroimidazo[4,5-b]indole derivatives. The envisaged methodology involves the L-proline-catalyzed condensation of para-amino-functionalized azo benzene, indoline-2,3-dione, and ammonium acetate precursors with pertinent aryl aldehyde derivatives under ultrasonic conditions. The structures of synthesized compounds were corroborated through FT-IR, 1H NMR, 13C NMR, and mass analysis data. Molecular docking studies assessed the inhibitory potential of these compounds against the main protease (Mpro) of SARS-CoV-2. Remarkably, in silico investigations revealed significant inhibitory action surpassing standard drugs such as Remdesivir, Paxlovid, Molnupiravir, Chloroquine, Hydroxychloroquine (HCQ), and (N3), an irreversible Michael acceptor inhibitor. Furthermore, the highly active compound was also screened for cytotoxicity activity against HEK-293 cells and exhibited minimal toxicity across a range of concentrations, affirming its favorable safety profile and potential suitability. The pharmacokinetic properties (ADME) of the synthesized compounds have also been deliberated. This study paves the way for in vitro and in vivo testing of these scaffolds in the ongoing battle against SARS-CoV-2.

An eco-friendly and cost-effective HPTLC method for quantification of COVID-19 antiviral drug and co-administered medications in spiked human plasma.

The coronavirus-2 has led to a global pandemic of COVID-19 with an outbreak of severe acute respiratory syndrome leading to worldwide quarantine measures and a rise in death rates. The objective of this study is to propose a green, sensitive, and selective densitometric method to simultaneously quantify remdesivir (REM) in the presence of the co-administered drug linezolid (LNZ) and rivaroxaban (RIV) in spiked human plasma. TLC silica gel aluminum plates 60 F254 were used as the stationary phase, and the mobile phase was composed of dichloromethane (DCM): acetone (8.5:1.5, v/v) with densitometric detection at 254 nm. Well-resolved peaks have been observed with retardation factors (Rf) of 0.23, 0.53, and 0.72 for REM, LNZ, and RIV, respectively. A validation study was conducted according to ICH Q2 (R1) Guidelines. The method was rectilinear over the concentration ranges of 0.2-5.5 µg/band, 0.2-4.5 µg/band and 0.1-3.0 µg/band for REM, LNZ and RIV, respectively. The sensitivities of REM, LIN, and RIV were outstanding, with quantitation limits of 128.8, 50.5, and 55.8 ng/band, respectively. The approach has shown outstanding recoveries ranging from 98.3 to 101.2% when applied to pharmaceutical formulations and spiked human plasma. The method's greenness was assessed using Analytical Eco-scale, GAPI, and AGREE metrics.

Molnupiravir increases SARS-CoV-2 genome diversity and complexity: A case-control cohort study.

Molnupiravir, an oral direct-acting antiviral effective in vitro against SARS-CoV-2, has been largely employed during the COVID-19 pandemic, since December 2021. After marketing and widespread usage, a progressive increase in SARS-CoV-2 lineages characterized by a higher transition/transversion ratio, a characteristic signature of molnupiravir action, appeared in the Global Initiative on Sharing All Influenza Data (GISAID) and International Nucleotide Sequence Database Collaboration (INSDC) databases. Here, we assessed the drug effects by SARS-CoV-2 whole-genome sequencing on 38 molnupiravir-treated persistently positive COVID-19 outpatients tested before and after treatment. Seventeen tixagevimab/cilgavimab-treated outpatients served as controls. Mutational analyses confirmed that SARS-CoV-2 exhibits an increased transition/transversion ratio seven days after initiation of molnupiravir. Moreover we observed an increased G->A ratio compared to controls, which was not related to apolipoprotein B mRNAediting enzyme, catalytic polypeptide-like (APOBEC) activity. In addition, we demonstrated for the first time an increased diversity and complexity of the viral quasispecies.

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.

Inhibition Mechanism of SARS-CoV-2 Infection by a Cholesterol Derivative, Nat-20(S)-yne.

The coronavirus disease 2019 (COVID-19) is caused by the etiological agent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19, with the recurrent epidemics of new variants of SARS-CoV-2, remains a global public health problem, and new antivirals are still required. Some cholesterol derivatives, such as 25-hydroxycholesterol, are known to have antiviral activity against a wide range of enveloped and non-enveloped viruses, including SARS-CoV-2. At the entry step of SARS-CoV-2 infection, the viral envelope fuses with the host membrane dependent of viral spike (S) glycoproteins. From the screening of cholesterol derivatives, we found a new compound 26,27-dinorcholest-5-en-24-yne-3ß,20-diol (Nat-20(S)-yne) that inhibited the SARS-CoV-2 S protein-dependent membrane fusion in a syncytium formation assay. Nat-20(S)-yne exhibited the inhibitory activities of SARS-CoV-2 pseudovirus entry and intact SARS-CoV-2 infection in a dose-dependent manner. Among the variants of SARS-CoV-2, inhibition of infection by Nat-20(S)-yne was stronger in delta and Wuhan strains, which predominantly invade into cells via fusion at the plasma membrane, than in omicron strains. The interaction between receptor-binding domain of S proteins and host receptor ACE2 was not affected by Nat-20(S)-yne. Unlike 25-hydroxycholesterol, which regulates various steps of cholesterol metabolism, Nat-20(S)-yne inhibited only de novo cholesterol biosynthesis. As a result, plasma membrane cholesterol content was substantially decreased in Nat-20(S)-yne-treated cells, leading to inhibition of SARS-CoV-2 infection. Nat-20(S)-yne having a new mechanism of action may be a potential therapeutic candidate for COVID-19.

SARS-CoV-2 aberrantly elevates mitochondrial bioenergetics to induce robust virus propagation.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a 'highly transmissible respiratory pathogen, leading to severe multi-organ damage. However, knowledge regarding SARS-CoV-2-induced cellular alterations is limited. In this study, we report that SARS-CoV-2 aberrantly elevates mitochondrial bioenergetics and activates the EGFR-mediated cell survival signal cascade during the early stage of viral infection. SARS-CoV-2 causes an increase in mitochondrial transmembrane potential via the SARS-CoV-2 RNA-nucleocapsid cluster, thereby abnormally promoting mitochondrial elongation and the OXPHOS process, followed by enhancing ATP production. Furthermore, SARS-CoV-2 activates the EGFR signal cascade and subsequently induces mitochondrial EGFR trafficking, contributing to abnormal OXPHOS process and viral propagation. Approved EGFR inhibitors remarkably reduce SARS-CoV-2 propagation, among which vandetanib exhibits the highest antiviral efficacy. Treatment of SARS-CoV-2-infected cells with vandetanib decreases SARS-CoV-2-induced EGFR trafficking to the mitochondria and restores SARS-CoV-2-induced aberrant elevation in OXPHOS process and ATP generation, thereby resulting in the reduction of SARS-CoV-2 propagation. Furthermore, oral administration of vandetanib to SARS-CoV-2-infected hACE2 transgenic mice reduces SARS-CoV-2 propagation in lung tissue and mitigates SARS-CoV-2-induced lung inflammation. Vandetanib also exhibits potent antiviral activity against various SARS-CoV-2 variants of concern, including alpha, beta, delta and omicron, in in vitro cell culture experiments. Taken together, our findings provide novel insight into SARS-CoV-2-induced alterations in mitochondrial dynamics and EGFR trafficking during the early stage of viral infection and their roles in robust SARS-CoV-2 propagation, suggesting that EGFR is an attractive host target for combating COVID-19.

In-vitro and in-vivo assessment of nirmatrelvir penetration into CSF, central nervous system cells, tissues, and peripheral blood mononuclear cells.

Three years after SARS-CoV-2 emerged as a global infectious threat, the virus has become endemic. The neurological complications such as depression, anxiety, and other CNS complications after COVID-19 disease are increasing. The brain, and CSF have been shown as viral reservoirs for SARS-CoV-2, yielding a potential hypothesis for CNS effects. Thus, we investigated the CNS pharmacology of orally dosed nirmatrelvir/ritonavir (NMR/RTV). Using both an in vitro and an in vivo rodent model, we investigated CNS penetration and potential pharmacodynamic activity of NMR. Through pharmacokinetic modeling, we estimated the median CSF penetration of NMR to be low at 18.11% of plasma with very low accumulation in rodent brain tissue. Based on the multiples of the 90% maximal effective concentration (EC90) for SARS-CoV-2, NMR concentrations in the CSF and brain do not achieve an exposure level similar to that of plasma. A median of only 16% of all the predicted CSF concentrations in rats were > 3xEC90 (unadjusted for protein binding). This may have implications for viral persistence and neurologic post-acute sequelae of COVID-19 if increased NMR penetration in the CNS leads to decreased CNS viral loads and decreased CNS inflammation.

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.

Deciphering the similarities and disparities of molecular mechanisms behind respiratory epithelium response to HCoV-229E and SARS-CoV-2 and drug repurposing, a systems biology approach.

PURPOSE: Identifying the molecular mechanisms behind SARS-CoV-2 disparities and similarities will help find new treatments. The present study determines networks' shared and non-shared (specific) crucial elements in response to HCoV-229E and SARS-CoV-2 viruses to recommend candidate medications. METHODS: We retrieved the omics data on respiratory cells infected with HCoV-229E and SARS-CoV-2, constructed PPIN and GRN, and detected clusters and motifs. Using a drug-gene interaction network, we determined the similarities and disparities of mechanisms behind their host response and drug-repurposed. RESULTS: CXCL1, KLHL21, SMAD3, HIF1A, and STAT1 were the shared DEGs between both viruses' protein-protein interaction network (PPIN) and gene regulatory network (GRN). The NPM1 was a specific critical node for HCoV-229E and was a Hub-Bottleneck shared between PPI and GRN in HCoV-229E. The HLA-F, ADCY5, TRIM14, RPF1, and FGA were the seed proteins in subnetworks of the SARS-CoV-2 PPI network, and HSPA1A and RPL26 proteins were the seed in subnetworks of the PPI network of HCOV-229E. TRIM14, STAT2, and HLA-F played the same role for SARS-CoV-2. Top enriched KEGG pathways included cell cycle and proteasome in HCoV-229E and RIG-I-like receptor, Chemokine, Cytokine-cytokine, NOD-like receptor, and TNF signaling pathways in SARS-CoV-2. We suggest some candidate medications for COVID-19 patient lungs, including Noscapine, Isoetharine mesylate, Cycloserine, Ethamsylate, Cetylpyridinium, Tretinoin, Ixazomib, Vorinostat, Venetoclax, Vorinostat, Ixazomib, Venetoclax, and epoetin alfa for further in-vitro and in-vivo investigations. CONCLUSION: We suggested CXCL1, KLHL21, SMAD3, HIF1A, and STAT1, ADCY5, TRIM14, RPF1, and FGA, STAT2, and HLA-F as critical genes and Cetylpyridinium, Cycloserine, Noscapine, Ethamsylate, Epoetin alfa, Isoetharine mesylate, Ribavirin, and Tretinoin drugs to study further their importance in treating COVID-19 lung complications.