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Background: The rapid emergence of the SARS-CoV-2 Omicron variant that evades many therapies illustrates the need for antiviral treatments with high genetic barriers to resistance. The small molecule PAV-104, identified through a moderate-throughput screen involving cell-free protein synthesis, was recently shown to target a subset of host protein assembly machinery in a manner specific to viral assembly with minimal host toxicity. The chemotype shows broad activity against respiratory viral pathogens, including Orthomyxoviridae, Paramyxoviridae, Adenoviridae, Herpesviridae, and Picornaviridae, with low susceptibility to evolutionary escape. Here, we investigated the capacity of PAV-104 to inhibit SARS-CoV-2 replication in human airway epithelial cells (AECs). Method(s): Dose-dependent cytotoxicity of PAV-104 in Calu-3 cells was determined by MTT assay. Calu-3 cells were infected with SARS-CoV-2 isolate USA-WA1/2020 (MOI=0.01). Primary AECs were isolated from healthy donor lung transplant tissue, cultured at air liquid interface (ALI), and infected with SARS-CoV-2 Gamma, Delta, and Omicron variants (MOI=0.1). SARS-CoV-2 replication was assessed by RT-PCR quantitation of the N gene, immunofluorescence assay (IFA) of nucleocapsid (N) protein, and titration of supernatant (TCID50). Transient co-expression of four SARS-CoV-2 structural proteins (N, M, S, E) to produce virus-like particles (VLPs) was used to study the effect of PAV-104 on viral assembly. Drug resin affinity chromatography was performed to study the interaction between PAV-104 and N. Glycerol gradient sedimentation was used to assess N oligomerization. Total RNA-seq and the REACTOME database were used to evaluate PAV-104 effects on the host transcriptome. Result(s): PAV-104 reached 50% cytotoxicity in Calu-3 cells at 3732 nM (Fig.1A). 50 nM PAV-104 inhibited >99% of SARS-CoV-2 infection in Calu-3 cells (p< 0.01) and in primary AECs (p< 0.01) (Fig.1B-E). PAV-104 specifically inhibited SARS-CoV-2 post entry, and suppressed production of SARS-CoV-2 VLPs without affecting viral protein synthesis. PAV-104 interacted with SARS-CoV-2 N and interfered with N oligomerization. Transcriptome analysis revealed that PAV-104 treatment reversed SARS-CoV-2 induction of the interferon and maturation of nucleoprotein signaling pathways. Conclusion(s): PAV-104 is a pan-respiratory virus small molecule inhibitor with promising activity against SARS-CoV-2 in human airway epithelial cells that should be explored in animal models and clinical studies.
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The regulation of inflammation is a complex pathophysiological process that depends on the production of oxygenated lipid derivatives essential polyunsaturated fatty acids, like omega-3 and omega-6, among which are the lipoxins resolvins and protectins, called specialized pro-resolving lipid mediators (SPM). Their activity is associated with the control of respiratory infection processes to modulate the production of proinflammatory cytokines, avoiding damage due to inflammation-associated necrosis, reducing microbial loads, and promoting tissue remodeling. Therefore, we review some of the biochemical, physiological and immunological aspects of SPM in the regulation of inflammation in respiratory infections.Copyright © 2022, Instituto Nacional de Enfermedades Respiratorias. All rights reserved.
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Biomimetic strategies can be adopted to improve biopharmaceutical aspects. Subsequently, Biomimetic reconstitutable pegylated amphiphilic lipid nanocarriers have high translational potential for systemic controlled drug delivery;however, such an improvised system for systemic aspirin delivery exploring nanotechnology is not available. Systemic administration of aspirin and its controlled delivery can significantly control blood clotting events, leading to stroke, which has immediate applications in cardiovascular diseases and Covid-19. In this work, we are developing aspirin sustained release pegylated amphiphilic self-assembling nanoparticles to develop reconstitutable aspirin injections by solvent-based co-precipitation method with phase inversion technique that leads to novel "biomimetic niosomal nanoparticles (BNNs).” DOE led optimization is done to develop Design of space for optimized particles. Upon reconstitution of solid powder, the particle size was 144.8 ± 12.90 nm with a surface charge of -29.2 ± 2.24 mV. The entrapment efficiency was found to be 49 ± 0.15%, wherein 96.99 ± 1.57% of the drug was released in 24hr showing super case II transport-based drug release mechanism. The formulation has the least hemolysis while showing significant suppression of platelet aggregation. MTT assay does not show any significant cytotoxicity. This is a potential nanoparticle that can be explored for developing aspirin injection, which is not available.
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The ongoing pandemic caused by the novel coronavirus, SARS-CoV-2, is influencing global health. Moreover, there is a major threat of future coronaviruses affecting the entire world in a similar, or even more dreadful, manner. Therefore, effective and biocompatible therapeutic options against coronaviruses are urgently needed. To address this challenge, medical specialists require a well-informed and safe approach to treating human coronaviruses (HCoVs). Herein, an environmental friendly approach for viral inactivation, based on plasma technology, was considered. A microwave plasma system was employed for the generation of the high amount of gaseous nitric oxide to prepare nitric oxide enriched plasma-activated water (NO-PAW), the effects of which on coronaviruses, have not been reported to date. To determine these effects, alpha-HCoV-229E was used in an experimental model. We found that NO-PAW treatment effectively inhibited coronavirus infection in host lung cells, visualized by evaluating the cytopathic effect and expression level of spike proteins. Interestingly, NO-PAW showed minimal toxicity towards lung host cells, suggesting its potential for therapeutic application. Moreover, this new approach resulted in viral inactivation and greatly improved the gene levels involved in host antiviral responses. Together, our findings provide evidence of an initiation point for further progress toward the clinical development of antiviral treatments, including such coronaviruses. © 2022 The Authors
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Introduction: COVID-19 is an acute respiratory tract disease caused by the emerging coronavirus SARS-CoV-2. Although several options for che-moprophylaxis are under development, effective treatment for COVID-19 is not yet available. Objective(s): To investigate the antiviral properties of synthesized silver na-noparticles (AgNPs) against SARS-CoV-2 using in vitro models. Material(s) and Method(s): This work synthesized AgNPs using an electrochemical method and characterized them using physico-chemical techniques (ICP-OES, ultraviolet-visible spectroscopy, and transmission electron microscopy). AgNPs with diameter sizes ranging between 2.6 to 30 nm and an average size of 6.2 nm were obtained by the electrochemical method. The cytotoxic effect and the antiviral activity of prepared AgNPs against SARS-CoV-2 were evaluated in vitro using Vero E6 cells. Cell viability was evaluated by MTT assay in the presence of serial dilutions of AgNPs. The antiviral effect of AgNPs was evaluated before and after the infection of Vero E6 cells by plaque assay. Result(s): Cytotoxic effect was observed at concentrations above 0.07 ppm. AgNPs exhibit a significant reduction of SARS-CoV-2 viral titer after a pre-post treatment strategy with inhibition of 96.5%, 64.13%, and 74.72% at 0.03, 0.017, and 0.008 ppm, respectively. Conclusion(s): Our results suggest that AgNPs could reduce SARS-CoV-2 replication with a low cytotoxic effect. Still, additional in vitro and in vivo studies are required to define its potential therapeutic application in humans. Copyright © 2023 Universidad de Antioquia.
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Background: Mono(ethylhexyl) phthalate (MEHP) is the main metabolite of Di(2-ethylhexyl) phthalate (DEHP), a chemical worldwide used as a plastic softener to increase the malleability, flexibility, and durability of several types of plastic, including those employed in bottled water, medical devices, and food wrapping, among others. Importantly, the consumption of these products has dramatically increased during the COVID-19 pandemic. MEHP has been classified as an endocrine disruptor chemical (EDC) and its involuntary intake has been associated with pregnancy complications such as preeclampsia and miscarriages. The placenta is a transitory organ that provides sustainability to the fetus, as well as the transportation of nutrients, hormones, and oxygen. Recent studies have proposed that MEHP may impair placental development and functionality. Nevertheless, little is known about its molecular mechanisms and effects on the placenta. Recent data has suggested that Sirtuin 1 (SIRT1) might be a molecular target. The aim of this study was to analyze the toxic and transcriptomic effects of MEHP in the human trophoblastic cell line HTR-8/svneo, focusing on the SIRT1-related pathways. Methods and Results: The HTR-8/svneo cell line was used as an extravillous trophoblast model to investigate MEHP effects. MEHP concentrations employed in this study were 0.5, 5, 50, 100, and 200 microM. Cell viability was evaluated by two methods: fixable viability staining, using eFluor 780, and MTT assay. Only the MTT assay suggested a significant decrease in cell viability at 48 hours with MEHP treatments of 5, 50, 100, and 200 microM. Mitochondrial biogenesis was analyzed by qPCR amplifying a region of the MT-ND1 mitochondrial gene. GAPDH promoter was used as a reference control. The results suggested a decrease in mitochondrial DNA at 48 hours. The transcriptomic analysis was performed in an Illumina Next-seq 500 with a coverage of 10 million reads. Doses of 5 and 200 microM of MEHP at 48 hours were analyzed. The results show that 41 and 341 genes were differentially expressed, respectively. These genes are involved in trophoblast function and pathophysiology and, according to previous reports, a portion of them are regulated by SIRT1. Finally, the effect of MEHP on SIRT1 was explored at both protein and mRNA levels by western blot and RT-qPCR, respectively. The results for mRNA levels exhibited a significant decline at 24 hours for all treatments, while protein levels were significantly reduced by 200 microM MEHP treatment at 48 hours. Conclusion(s): The present study demonstrates that MEHP treatments promote mitochondrial dysfunction in HTR-8/svneo cells. Moreover, the transcriptome analysis showed that MEHP modifies important signals for placental function and pathophysiology. The decline in SIRT1 levels correlates with the mitochondrial alterations as well as a portion of the transcriptomic changes, suggesting that SIRT1 may have an important role in MEHP effects in trophoblastic cells. Copyright © 2022 Elsevier B.V.
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A series of 4-Aminophenyl-1H 1,2,3-traizole based benzofuran analogs were synthesized with high yields straightforwardly via microwave irradiation methods than conventional methods. The structures of synthesized compounds were confirmed based on IR, 1H, 13C NMR, and HR mass spectral analysis. All the synthesized compounds were subjected to evaluation of their in vitro anticancer activity with MCF-7, PC-3, and HeLa cell lines by MTT assay, and compounds 7m, 7j, 7e, 7g, 7b, 7h, and 7c have shown good results compared to standard Doxorubicin. Further, in vitro antimicrobial activity of synthesized compounds examined by Agar Disc Diffusion method by taking gram-positive, gram-negative bacterial, and fungal strains and turned out with encouraging results compared to standard Ciprofloxacin and Fluconazole respectively. Furthermore, molecular docking studies were carried out to find out H-bonding interactions, hydrophobic interaction with docking scores of synthesized compounds. We have carried out molecular docking on COVID-19 study with COVID-19 main protease enzyme and got outstanding binding interactions. © 2022
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Covid-19 was mainly treated by a broad-spectrum antiviral called Remdesivir. A truncated cone molecular structure of Hydroxypropyl-ß-cyclodextrin can enhance the solubility and cellular uptake of the poorly soluble drug's through biological membranes. This study aimed to synthesize, characterize, observe cellular uptake and evaluate the cytotoxicity of remdesivirhydroxypropyl-ß-cyclodextrin (RDV-HPßCD) inclusion complex. The RDV-HPßCD inclusion complex was synthesized by the solvent evaporation method. Furthermore, the inclusion complex characteristic was evaluated by ultraviolet-visible (UV-Vis) spectrophotometry;particle size analyzer (PSA);Fourier infrared spectrophotometry (FTIR);X-ray diffraction (XRD);and differential scanning calorimetry (DSC). Further, fluorescence microscopy was used to evaluate the cellular uptake and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used in the cytotoxicity study. In the UV-Vis spectrum, both the inclusion complex and pure remdesivir showed a maximum peak at 246 nm. The inclusion complex has a particle size of 1697 ± 738.02 nm with -22.4 ± 1.58 mV of zeta potential. Shifted FTIR spectrum, broad XRD peak, and broad DSC thermogram peak at 72.93 °C indicated the successful formation of the RDV-HPßCD inclusion complex. Furthermore, cellular uptake observation of RDV-HPßCD inclusion complex conjugated to FITC showed better intensity inside the Vero cell than pure remdesivir conjugated to FITC. Further, Inclusion complex showed higher cell viability than pure remdesivir at a certain concentration.
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Introduction: The rapid emergence of the SARS-CoV-2 Omicron variant that evades many monoclonal antibody therapies illustrates the need for anti-viral treatments with low susceptibility to evolutionary escape. The small molecule PAV-104, identified through a moderate-throughput screen involving cell-free protein synthesis, was recently shown to target a subset of host protein assembly machinery in a manner specific to viral assembly. This compound has minimal host toxicity, including once daily oral dosing in rats that achieves >200-fold of the 90% effective concentration (EC90) in blood. The chemotype shows broad activity against respiratory viral pathogens, including Orthomyxoviridae, Paramyxoviridae, Adenoviridae, Herpesviridae, and Picornaviridae, with low suceptability to evolutionary escape. We hypothesized that PAV-104 would be active against SARSCoV- 2 variants in human airway epithelial cells. Methods: Airway epithelial cells were differentiated from lung transplant tissue at air-liquid interface (ALI) for four weeks prior to challenge with Alpha (Pango lineage designation B.1.1.7), Beta (B.1.351), Gamma (P.1), and Delta (B.1.617.2) SARS-CoV-2 variants. Viral replication was determined by quantitative PCR measurement of the SARS-CoV-2 nucleocapsid (N) gene. Dose-dependent virus inhibition and cytotoxicity of PAV-104 in the Calu-3 airway epithelial cell line was determined by PCR and MTT assay. Student's t-tests were used to evaluate statistical significance. Results: Alpha, Beta, Gamma, and Delta variants of SARS-CoV-2 showed comparable infectivity in human primary airway epithelial cells at ALI (N=3 donors), 47- to 550-fold higher than the parent (USA-WA1/2020) strain. PAV-104 reached 50% cytotoxicity in Calu-3 cells at 240 nM (Fig. 1A). Dose-response studies in Calu-3 cells demonstrated PAV-104 has a 6 nM 50% inhibitory concentration (IC50) for blocking replication of SARS-CoV-2 (USA-WA1/2020) (Fig.1B). In primary cells at ALI from 3 donors tested, there was >99% inhibition of infection by SARS-CoV-2 Gamma variant (N=3, MOI 0.1, P <0.01) with 100 nM PAV-104 (Fig. 1C). Addition of 100 nM PAV-104 2-hours post-infection, but not pre-infection, resulted in >99% suppression of viral replication, indicating a post-entry drug mechanism. PAV-104 bound a small subset of the known allosteric modulator 14-3-3, itself implicated in the interactome of SARS-CoV-2. Conclusion: PAV-104 is a host-targeted, orally bioavailable, pan-viral small molecule inhibitor with promising activity against SARS-CoV-2 variants in human primary airway epithelial cells. (Figure Presented).
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Background: Galectin-9 (Gal-9) is a β-galactoside-binding lectin involved in immune regulation and viral immunopathogenesis. Multiple recent reports demonstrate that plasma levels of Gal-9 are elevated in the setting of severe COVID-19 disease. However, a causal role of Gal-9 in SARS-CoV-2 pathology remains to be elucidated. Here, we determined the impact of Gal-9 on SARS-CoV-2 replication and pro-inflammatory signaling in immortalized and primary human airway epithelial cells (AECs). Methods: Dose-dependent cytotoxicity of recombinant human Gal-9 in the Calu-3 AEC line was determined by MTT assay. Calu-3 cells were infected with SARS-CoV-2 isolate USA-WA1/2020 (MOI=0.01). Primary AECs were isolated from healthy donor lung transplant tissue, cultured at air liquid interface (ALI), and infected with SARS-CoV-2 lineage P.1 (MOI=0.1). SARS-CoV-2 replication was assessed by RT-PCR quantitation of the nucleocapsid (N) gene, immunofluorescence assay (IFA) of N protein, and titration of supernatant (TCID50). Viral entry was measured using luciferase activity of VSV-SARS-CoV-2 S-ΔG-Luciferase reporter pseudovirus. ACE2 and TMPRSS2 cell-surface expression were measured by flow cytometry. Pro-inflammatory factors (IL-6, IL-8, and TNFα) were detected by RT-PCR. Total RNA-seq was used to evaluate Gal-9 effects on the host transcriptome. Groups were compared by Student's t-test, and differential expression analyses were performed using DESeq2. Results: Gal-9 reached 50% cytotoxicity in Calu-3 cells at 597 nM. Gal-9 significantly increased SARS-CoV-2 expression (8.1 to 25.5 fold;p<0.0001) and infectious virus release (1.9 to 17.8 fold;p<0.038) in a dose-dependent manner in Calu-3 cells. Pseudovirus entry into Calu-3 cells was enhanced by Gal-9 (2.4 to 5.6 fold;p<0.0016), and the enhanced entry was inhibited by anti-ACE2 antibody (p<0.0027). Cell surface ACE2 and TMPRSS2 expression were unaffected by Gal-9. Gal-9 treatment accelerated virus-induced expression of IL-6, IL-8, and TNFα (p<0.018) in Calu-3 cells. Gal-9 increased SARS-CoV-2 production (p=0.03) and pro-inflammatory factor expression (p<0.05) in primary AECs (N=5 donors). RNA-seq data revealed that Gal-9 significantly induced IL-17, EIF2, IL-8 and IL-6 signaling pathways in the setting of SARS-CoV-2 infection. Conclusion: Gal-9 facilitates SARS-CoV-2 entry, replication, and virus-induced pro-inflammatory signaling in AECs ex vivo. Our data suggest that pharmacologic manipulation of Gal-9 should be explored as a SARS-CoV-2 therapeutic strategy.
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The ability to control the proliferation and cell death by inhibiting specific target kinase offers the opportunity to apply targeted therapies in the treatment of cancer. It has been found that (S)-valine-thiazole-derived compounds such as NEOS-223 are effective inhibitors of one or more of these kinases. NEOS 223 was developed, synthesized, and tested in the NCI 60 human tumor cell-screening panel demonstrating inhibition of colon (-53%), melanoma (-41%), and breast cancers (-9%). Microsomal clearance was determined in mouse, rat, dog, and human, and analyzed by LC-MS/MS by percent of parent material. IC50 values for CYP inhibition of >10 μM were calculated for 1A2, 2C19, and 3A4 with IC50 values of 4.86, 4.31, and 7.84 μM for 2C9 and 2D6. Microsomal clearance was high in all species with clearance rates ranging from 69-136 mL/min/kg. Plasma protein binding was determined by Rapid Equilibrium Dialysis in mice, rats, dogs, and humans. High plasma protein binding (>70%) was observed across all species. Based on the NCI results several cell lines were assayed in an MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide) to determine cell viability in the presence of NEOS-223 resulting in <20% viability in colon, breast, melanoma, pancreatic and prostate human cancer cell lines at a 10 uM concentration. Maximum tolerated dose studies were conducted by both intraperitoneal and oral administration in mice. NEOS-223 delivered up to 80 mg/kg was well tolerated. Minimal or no toxicity was observed in acute and repeat dose animal studies. Pharmacokinetics of oral administration demonstrated adequate systemic exposure at therapeutic levels in mice, rats, and dogs. Preliminary in vivo mouse xenograft studies were performed on colon (COLO 205, HT-29 red FLUC), breast (MDA-Sumathi Chittamuru;Timothy M. Murphy;Sara A. Little;Andrew A. Taylor;Roseanne Wexler;Laxman Desai MB-468), melanoma (M-14), pancreatic (PANC-1), and prostate (PC3) human cancer cells with significant tumor inhibition observed compared to positive control agent groups with twice daily dosing of NEOS-223. In addition, a five-day pilot oral toxicity study in rats with dose range-finding studies and a 28-day repeat dose toxicity study performed in both rats and dogs provided favorable results. NEOS-223 has demonstrated active in vitro activity along with a favorable safety profile. in vivo efficacy resulted in inhibition of growth of multiple cell line. As a novel effective structure possibly targeting multiple kinases and transporters in one hybrid molecule, NEOS-223 may be a preferred monotherapy or combined therapy for multiple cancers. If upon further development, this drug is effective in humans, it would advance clinical practice and could improve current therapy significantly.
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The diversity in Jordan’s flora due to its geographical areas make is well noted in the scientific literature. The challenge of disease and death caused by infectious diseases like viruses and bacteria, and as infectious diseases evolve and pathogens develop resistance to existing pharmaceuticals, the search for new novel leads, possibly with different modes of action, against bacterial and viral diseases has intensified in recent years. The intent of this review is to provide prevalent information on the antibacterial and antiviral potential in medicinal plants in Jordan, mode of action, type of viruses and bacteria, and phytochemical contents. It has been demonstrated by several studies presented in this review that medicinal plants in Jordan are rich in phytochemicals and possess antiviral and antibacterial properties.
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Azithromycin (AZM) is a macrolide antibiotic used for the treatment of various bacterial infections. The drug is known to have low oral bioavailability (37%) which may be attributed to its relatively high molecular weight, low solubility, dissolution rate, and incomplete intestinal absorption. To overcome these drawbacks, liquid (L) and solid (S) self-emulsifying drug delivery systems (SEDDs) of AZM were developed and optimized. Eight different pseudo-ternary diagrams were constructed based on the drug solubility and the emulsification studies in various SEDDs excipients at different surfactant to co-surfactant (Smix) ratios. Droplet size (DS) < 150 nm, dispersity (D) ≤ 0.7, and transmittance (T)% > 85 in three diluents of distilled water (DW), 0.1 mM HCl, and simulated intestinal fluids (SIF) were considered as the selection criteria. The final formulations of L-SEDDs (L-F1(H)), and S-SEDDs (S-F1(H)) were able to meet the selection requirements. Both formulations were proven to be cytocompatible and able to open up the cellular epithelial tight junctions (TJ). The drug dissolution studies showed that after 5 min > 90% and 52.22% of the AZM was released from liquid and solid SEDDs formulations in DW, respectively, compared to 11.27% of the pure AZM, suggesting the developed SEDDs may enhance the oral delivery of the drug. The formulations were stable at refrigerator storage conditions.