RESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Morus alba L. are widely used as ethnomedicine and functional food in China, Japan, Korea and other Asian countries. Morus alba L. have a variety of pharmacological activity such as antiviral, antioxidation, anti-cholesterol, anticancer, hypoglycemia, and neuroprotection. Morus alba L. has demonstrated antiviral efficacy against influenza viruses, SARS-CoV-2 and so on, but its potential activity against pseudorabies virus (PRV) remains uncertain. AIM OF THE STUDY: This study endeavors to delve into the anti-pseudorabies virus (PRV) potential of the ethanol extract of Morus alba L. leaves (MLE), while simultaneously elucidating its underlying mechanism of action. MATERIALS AND METHODS: The anti-PRV activities of Morus alba L. extracts at different concentrations were evaluated by qPCR and immunoblotting. The inhibitory effects of MLE on PRV replication in three distinct treatment modes (pretreatment, co-treatment, and post-treatment) were detected by qPCR and indirect immunofluorescence assays. qPCR was used to investigate the effects of MLE on PRV attachment, entrance, and cytokine expression in PRV-infected cells. The chemical components in MLE were analyzed by UPLC-MS/MS. RESULTS: MLE significantly inhibits PRV replication and protein expression in a dose-dependent manner. MLE displays inhibitory effects against PRV at three different modes of treatment. The most significant inhibitory effect of MLE was observed when used in co-treatment mode, resulting in an inhibition rate of 99.42%. MLE inhibits PRV infection in the early stage. MLE inhibits PRV infection by affecting viral attachment and viral entry. Furthermore, MLE exerts its inhibition on PRV replication by mitigating the heightened expression of cytokines (TNF-α and IFN-α) triggered by PRV. Analysis of its chemical composition highlights phenolic acids and flavonoids as the principal constituents of MLE. CONCLUSION: The results illustrate that MLE effectively impedes PRV infection by suppressing viral adsorption and entry, while also curbing the expression of antiviral cytokines. Therefore, MLE may be a potential resource for creating new medications to treat human and animal PRV infections.
Assuntos
Antivirais , Herpesvirus Suídeo 1 , Morus , Extratos Vegetais , Folhas de Planta , Replicação Viral , Herpesvirus Suídeo 1/efeitos dos fármacos , Morus/química , Antivirais/farmacologia , Antivirais/isolamento & purificação , Extratos Vegetais/farmacologia , Animais , Replicação Viral/efeitos dos fármacos , Folhas de Planta/química , Citocinas/metabolismo , Cães , Células Madin Darby de Rim Canino , Internalização do Vírus/efeitos dos fármacos , Ligação Viral/efeitos dos fármacosRESUMO
The main vectors of Zika virus (ZIKV) and dengue virus (DENV) are Aedes aegypti and Ae. albopictus, with Ae. aegypti being more competent. However, the underlying mechanisms remain unclear. Here, we find Ae. albopictus shows comparable vector competence to ZIKV/DENV with Ae. aegypti by blood-feeding after antibiotic treatment or intrathoracic injection. This suggests that midgut microbiota can influence vector competence. Enterobacter hormaechei_B17 (Eh_B17) is isolated from field-collected Ae. albopictus and conferred resistance to ZIKV/DENV infection in Ae. aegypti after gut-transplantation. Sphingosine, a metabolite secreted by Eh_B17, effectively suppresses ZIKV infection in both Ae. aegypti and cell cultures by blocking viral entry during the fusion step, with an IC50 of approximately 10 µM. A field survey reveals that Eh_B17 preferentially colonizes Ae. albopictus compared to Ae. aegypti. And field Ae. albopictus positive for Eh_B17 are more resistant to ZIKV infection. These findings underscore the potential of gut symbiotic bacteria, such as Eh_B17, to modulate the arbovirus vector competence of Aedes mosquitoes. As a natural antiviral agent, Eh_B17 holds promise as a potential candidate for blocking ZIKV/DENV transmission.
Assuntos
Aedes , Vírus da Dengue , Enterobacter , Microbioma Gastrointestinal , Mosquitos Vetores , Esfingosina , Simbiose , Zika virus , Aedes/virologia , Aedes/microbiologia , Aedes/efeitos dos fármacos , Animais , Mosquitos Vetores/microbiologia , Mosquitos Vetores/virologia , Mosquitos Vetores/efeitos dos fármacos , Zika virus/fisiologia , Zika virus/efeitos dos fármacos , Vírus da Dengue/efeitos dos fármacos , Vírus da Dengue/fisiologia , Microbioma Gastrointestinal/efeitos dos fármacos , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Esfingosina/farmacologia , Enterobacter/efeitos dos fármacos , Enterobacter/fisiologia , Infecção por Zika virus/transmissão , Infecção por Zika virus/virologia , Dengue/transmissão , Dengue/virologia , Dengue/prevenção & controle , Feminino , Internalização do Vírus/efeitos dos fármacos , HumanosRESUMO
The life cycle of Ebola and Marburg viruses includes a step of the virion envelope fusion with the cell membrane. Here, we analyzed whether the fusion of liposome membranes under the action of fragments of fusion peptides of Ebola and Marburg viruses depends on the composition of lipid vesicles. A fluorescence assay and electron microscopy were used to quantify the fusogenic activity of the virus fusion peptides and to identify the lipid determinants affecting membrane merging. Differential scanning calorimetry of lipid phase transitions revealed alterations in the physical properties of the lipid matrix produced by virus fusion peptides. Additionally, we found that plant polyphenols, quercetin, and myricetin inhibited vesicle fusion induced by the Marburg virus fusion peptide.
Assuntos
Ebolavirus , Flavonoides , Marburgvirus , Ebolavirus/efeitos dos fármacos , Marburgvirus/efeitos dos fármacos , Marburgvirus/química , Flavonoides/química , Flavonoides/farmacologia , Fusão de Membrana/efeitos dos fármacos , Lipossomos/química , Quercetina/química , Quercetina/farmacologia , Internalização do Vírus/efeitos dos fármacos , Doença pelo Vírus Ebola/virologia , Polifenóis/química , Polifenóis/farmacologia , Proteínas Virais de Fusão/química , Proteínas Virais de Fusão/metabolismo , Humanos , Membrana Celular/metabolismo , Peptídeos/química , Peptídeos/farmacologia , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/farmacologiaRESUMO
MoMo30 is an antiviral protein isolated from aqueous extracts of Momordica balsamina L. (Senegalese bitter melon). Previously, we demonstrated MoMo30's antiviral activity against HIV-1. Here, we explore whether MoMo30 has antiviral activity against the COVID-19 virus, SARS-CoV-2. MLV particles pseudotyped with the SARS-CoV-2 Spike glycoprotein and a Luciferase reporter gene (SARS2-PsV) were developed from a three-way co-transfection of HEK293-T17 cells. MoMo30's inhibition of SARS2-PsV infection was measured using a luciferase assay and its cytotoxicity using an XTT assay. Additionally, MoMo30's interactions with the variants and domains of Spike were determined by ELISA. We show that MoMo30 inhibits SARS2-PsV infection. We also report evidence of the direct interaction of MoMo30 and SARS-CoV-2 Spike from WH-1, Alpha, Delta, and Omicron variants. Furthermore, MoMo30 interacts with both the S1 and S2 domains of Spike but not the receptor binding domain (RBD), suggesting that MoMo30 inhibits SARS-CoV-2 infection by inhibiting fusion of the virus and the host cell via interactions with Spike.
Assuntos
Antivirais , Ligação Proteica , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/química , Humanos , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/fisiologia , Células HEK293 , Antivirais/farmacologia , COVID-19/virologia , Internalização do Vírus/efeitos dos fármacos , Pseudotipagem ViralRESUMO
The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has filled a gap in our knowledge regarding the prevention of CoVs. Swine coronavirus (CoV) is a significant pathogen that causes huge economic losses to the global swine industry. Until now, anti-CoV prevention and control have been challenging due to the rapidly generated variants. Silver nanoparticles (AgNPs) with excellent antimicrobial activity have attracted great interest for biosafety prevention and control applications. In this study, we synthesized chitosan-modified AgNPs (Chi-AgNPs) with good biocompatibility to investigate their antiviral effects on swine CoVs. In vitro assays showed that Chi-AgNPs could significantly impaired viral entry. The direct interaction between Chi-AgNPs and CoVs can destroy the viral surface spike (S) protein secondary structure associated with viral membrane fusion, which is caused by the cleavage of disulfide bonds in the S protein. Moreover, the mechanism showed that Chi-AgNPs reduced the virus-induced apoptosis of Vero cells via the ROS/p53 signaling activation pathway. Our data suggest that Chi-AgNPs can serve as a preventive strategy for CoVs infection and provide a molecular basis for the viricidal effect of Chi-AgNPs on CoVs.
Assuntos
Antivirais , Quitosana , Nanopartículas Metálicas , Prata , Glicoproteína da Espícula de Coronavírus , Animais , Quitosana/química , Quitosana/farmacologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Nanopartículas Metálicas/química , Chlorocebus aethiops , Prata/química , Prata/farmacologia , Células Vero , Antivirais/farmacologia , Antivirais/química , Suínos , Apoptose/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , SARS-CoV-2/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , COVID-19/virologia , COVID-19/prevenção & controleRESUMO
The endemic nature of the Ebola virus disease in Africa underscores the need for prophylactic and therapeutic drugs that are affordable and easy to administer. Through a phenotypic screening employing viral pseudotypes and our in-house chemical library, we identified a promising hit featuring a thiophene scaffold, exhibiting antiviral activity in the micromolar range. Following up on this thiophene hit, a new series of compounds that retain the five-membered heterocyclic scaffold while modifying several substituents was synthesized. Initial screening using a pseudotype viral system and validation assays employing authentic Ebola virus demonstrated the potential of this new chemical class as viral entry inhibitors. Subsequent investigations elucidated the mechanism of action through site-directed mutagenesis. Furthermore, we conducted studies to assess the pharmacokinetic profile of selected compounds to confirm its pharmacological and therapeutic potential.
Assuntos
Antivirais , Barreira Hematoencefálica , Ebolavirus , Tiofenos , Internalização do Vírus , Tiofenos/química , Tiofenos/farmacocinética , Tiofenos/farmacologia , Tiofenos/síntese química , Ebolavirus/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Humanos , Antivirais/farmacologia , Antivirais/química , Antivirais/farmacocinética , Antivirais/síntese química , Internalização do Vírus/efeitos dos fármacos , Relação Estrutura-Atividade , Animais , Descoberta de Drogas , Administração Oral , Disponibilidade Biológica , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/virologiaRESUMO
Lactoferrin is a multifunctional glycoprotein naturally found in mammalian secretions, predominantly in colostrum and milk. As a key component of dairy foods, lactoferrin enhances viral protection and boosts human health, owing to its fundamental properties including antiviral, anti-inflammatory, and immune-modulatory effects. Importantly, the antiviral effect of lactoferrin has been shown against a range of viruses causing serious infections and threatening human health. One of the viruses that lactoferrin exerts significant antiviral effects on is the human papillomavirus (HPV), which is the most prevalent transmitted infection affecting a myriad of people around the world. Lactoferrin has a high potential to inhibit HPV via different mechanisms, including direct binding to viral envelope proteins or their cell receptors, thereby hindering viral entry and immune stimulation by triggering the release of some immune-related molecules through the body, such as lymphocytes. Along with HPV, lactoferrin also can inhibit a range of viruses including coronaviruses and hepatitis viruses in the same manner. Here, we overview the current knowledge of lactoferrin and its effects on HPV and other viral infections.
Assuntos
Antivirais , Lactoferrina , Infecções por Papillomavirus , Lactoferrina/uso terapêutico , Lactoferrina/farmacologia , Humanos , Infecções por Papillomavirus/tratamento farmacológico , Antivirais/farmacologia , Antivirais/uso terapêutico , Papillomaviridae/efeitos dos fármacos , Animais , Internalização do Vírus/efeitos dos fármacos , Papillomavirus HumanoRESUMO
Since combination antiretroviral therapy (cART) was introduced to treat human immunodeficiency virus type-1 (HIV-1)/acquired immunodeficiency syndrome (AIDS), the AIDS mortality rate has markedly decreased, and convalescence in individuals with HIV has improved drastically. However, sexual transmission has made HIV-1 a global epidemic. Sacran is a megamolecular polysaccharide extracted from cyanobacterium Aphanothece sacrum that exhibits numerous desirable characteristics for transdermic applications, such as safety as a biomaterial, a high moisture retention effect, the ability to form a film and hydrogel, and an anti-inflammatory effect. In this study, we evaluated the anti-HIV-1 effects in sacran as a barrier to HIV-1 transmission. Sacran inhibited HIV-1 infection and envelope-dependent cell-to-cell fusion. Moreover, we used a Transwell assay to confirm that sacran inhibited viral diffusion and captured viruses. The synergistic effects of sacran and other anti-HIV infection drugs were also evaluated. HIV-1 infections can be reduced through the synergistic effects of sacran and anti-HIV-1 drugs. Our study suggests using sacran gel to provide protection against HIV-1 transmission.
Assuntos
Infecções por HIV , HIV-1 , Humanos , HIV-1/efeitos dos fármacos , Infecções por HIV/prevenção & controle , Infecções por HIV/virologia , Cianobactérias/química , Fármacos Anti-HIV/farmacologia , Polissacarídeos Bacterianos/farmacologia , Polissacarídeos/farmacologia , Anti-Infecciosos/farmacologia , Internalização do Vírus/efeitos dos fármacos , Linhagem CelularRESUMO
Zika virus (ZIKV; family, Flaviviridae), which causes congenital Zika syndrome, Guillain-Barré Syndrome, and other severe diseases, is transmitted mainly by mosquitoes; however, the virus can be transmitted through other routes. Among the three structural and seven nonstructural proteins, the surface envelope (E) protein of ZIKV plays a critical role in viral entry and pathogenesis, making it a key target for the development of effective entry inhibitors. This review article describes the life cycle, genome, and encoded proteins of ZIKV, illustrates the structure and function of the ZIKV E protein, summarizes E protein-targeting entry inhibitors (with a focus on those based on natural products and small molecules), and highlights challenges that may potentially hinder the development of effective inhibitors of ZIKV infection. Overall, the article will provide useful guidance for further development of safe and potent ZIKV entry inhibitors targeting the viral E protein.
Assuntos
Antivirais , Proteínas do Envelope Viral , Internalização do Vírus , Infecção por Zika virus , Zika virus , Zika virus/efeitos dos fármacos , Zika virus/fisiologia , Internalização do Vírus/efeitos dos fármacos , Humanos , Proteínas do Envelope Viral/metabolismo , Proteínas do Envelope Viral/antagonistas & inibidores , Infecção por Zika virus/virologia , Infecção por Zika virus/tratamento farmacológico , Antivirais/farmacologia , AnimaisRESUMO
BACKGROUND: Pseudorabies virus (PRV), a member of the family Herpesviridae, is responsible for significant economic losses in the pig industry and has recently been associated with human viral encephalitis, leading to severe neurological symptoms post-recovery. Despite the widespread impact of PRV, there are currently no approved effective drugs for treating PRV-related diseases in humans or pigs. Therefore, the exploration and discovery of safe and effective drugs for the prevention and treatment of PRV infection is of paramount importance. PURPOSE: The objective of this study is to screen and identify natural compounds with antiviral activity against PRV. METHODS: First, we used a strain of PRV with green fluorescent protein (PRV-GFP) to screen a natural product chemical library to identify potential antiviral drugs. Next, we assessed the antiviral abilities of salvianolic acid A (SAA) in vitro using virus titer assay, qPCR, and IFA. We investigated the mechanisms of SAA's antiviral activity through viral attachment, internalization, inactivation, and nuclease digestion assay. Finally, we evaluated the efficacy of SAA in inactivating PRV using mice as the experimental subjects. RESULTS: This study screened 206 natural compounds for anti-PRV activity in vitro, resulting in the identification of seven potential antiviral agents. Notably, SAA emerged as a promising candidate with significant anti-PRV activity. The mechanism of action may be that SAA can directly inactivate the virus by disrupting viral envelope. In vivo experiments have shown that pre-incubation of SAA and PRV can effectively inhibit the infectivity and pathogenicity of PRV in mice. CONCLUSION: This study offers valuable insights into the antiviral properties of SAA, potentially informing strategies for controlling PRV epidemics and treating related diseases in both humans and animals.
Assuntos
Antivirais , Herpesvirus Suídeo 1 , Pseudorraiva , Herpesvirus Suídeo 1/efeitos dos fármacos , Animais , Antivirais/farmacologia , Pseudorraiva/tratamento farmacológico , Pseudorraiva/virologia , Camundongos , Ácidos Cafeicos/farmacologia , Lactatos/farmacologia , Internalização do Vírus/efeitos dos fármacos , Vírion/efeitos dos fármacos , Suínos , Ligação Viral/efeitos dos fármacos , Linhagem Celular , FemininoRESUMO
Cellular processes are controlled by the thermodynamics of the underlying biomolecular interactions. Frequently, structural investigations use one monomeric binding partner, while ensemble measurements of binding affinities generally yield one affinity representative of a 1:1 interaction, despite the majority of the proteome consisting of oligomeric proteins. For example, viral entry and inhibition in SARS-CoV-2 involve a trimeric spike surface protein, a dimeric angiotensin-converting enzyme 2 (ACE2) cell-surface receptor and dimeric antibodies. Here, we reveal that cooperativity correlates with infectivity and inhibition as opposed to 1:1 binding strength. We show that ACE2 oligomerizes spike more strongly for more infectious variants, while exhibiting weaker 1:1 affinity. Furthermore, we find that antibodies use induced oligomerization both as a primary inhibition mechanism and to enhance the effects of receptor-site blocking. Our results suggest that naive affinity measurements are poor predictors of potency, and introduce an antibody-based inhibition mechanism for oligomeric targets. More generally, they point toward a much broader role of induced oligomerization in controlling biomolecular interactions.
Assuntos
Enzima de Conversão de Angiotensina 2 , COVID-19 , Ligação Proteica , Multimerização Proteica , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , SARS-CoV-2/metabolismo , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Humanos , COVID-19/virologia , COVID-19/metabolismo , COVID-19/imunologia , Internalização do Vírus/efeitos dos fármacos , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/metabolismo , TermodinâmicaRESUMO
The white spot syndrome virus (WSSV), a rapidly replicating and highly lethal pathogen that targets Penaeid shrimp, has emerged as one of the most widespread viruses globally due to its high virulence. With effective chemotherapeutics still unavailable, the pursuit of novel and viable strategies against WSSV remains a crucial focus in the field of shrimp farming. The envelope proteins of WSSV are essential for virus entry, serving as excellent targets for the development of antiviral therapeutics. Novel strategies in the design of inhibitory peptides, especially those targeting envelope protein (VP28) located on the surface of the virus particle, play a critical role as a significant virulence factor during the early stages of inherent WSSV infection in shrimp. In this direction, the current computational study focused on identifying self-inhibitory peptides from the hydrophobic membrane regions of the VP28 protein, employing peptide docking and molecular dynamics simulation (MDS) approaches. Such inhibitory peptides could be useful building blocks for the rational engineering of inhibitory therapeutics since they imitate the mechanism of binding to homologous partners used by their origin domain to interact with other molecules. The N-terminal sequence of VP28 has been reported as the potential site for membrane interactions during the virus entry. Moreover, drug delivery systems mediated by chitosan and gold nanoparticles are being developed to enhance the therapeutic efficacy of anti-viral peptides. These systems can increase the solubility, stability, and selectivity of peptides, possessing better qualities than conventional delivery methods. This computational study on self-inhibitory peptides could be a valuable resource for further in vitro and in vivo studies on anti-viral therapeutics in the aquaculture industry.
Assuntos
Antivirais , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Penaeidae , Peptídeos , Vírus da Síndrome da Mancha Branca 1 , Vírus da Síndrome da Mancha Branca 1/efeitos dos fármacos , Vírus da Síndrome da Mancha Branca 1/genética , Antivirais/farmacologia , Animais , Peptídeos/farmacologia , Peptídeos/química , Penaeidae/virologia , Proteínas do Envelope Viral/metabolismo , Proteínas do Envelope Viral/química , Internalização do Vírus/efeitos dos fármacosRESUMO
Nipah virus (NiV) is known to be a highly pathogenic zoonotic virus, which is included in the World Health Organization Research & Development Blueprint list of priority diseases with up to 70% mortality rate. Due to its high pathogenicity and outbreak potency, a therapeutic countermeasure against NiV is urgently needed. As NiV needs to be handled within a Biological Safety Level (BSL) 4 facility, we had developed a safe drug screening platform utilizing a baculovirus expression vector system (BEVS) based on a NiV-induced syncytium formation that could be handled within a BSL-1 facility. To reconstruct the NiV-induced syncytium formation in BEVS, two baculoviruses were generated to express recombinant proteins that are responsible for inducing the syncytium formation, including one baculovirus exhibiting co-expressed NiV fusion protein (NiV-F) and NiV attachment glycoprotein (NiV-G) and another exhibiting human EphrinB2 protein. Interestingly, syncytium formation was observed in infected insect cells when the medium was modified to have a lower pH level and supplemented with cholesterol. Fusion inhibitory properties of several compounds, such as phytochemicals and a polysulfonated naphthylamine compound, were evaluated using this platform. Among these compounds, suramin showed the highest fusion inhibitory activity against NiV-induced syncytium in the baculovirus expression system. Moreover, our in silico results provide a molecular-level glimpse of suramin's interaction with NiV-G's central hole and EphrinB2's G-H loop, which could be the possible reason for its fusion inhibitory activity.
Assuntos
Baculoviridae , Avaliação Pré-Clínica de Medicamentos , Células Gigantes , Vírus Nipah , Vírus Nipah/genética , Vírus Nipah/efeitos dos fármacos , Baculoviridae/genética , Animais , Humanos , Células Gigantes/efeitos dos fármacos , Células Gigantes/metabolismo , Células Gigantes/virologia , Avaliação Pré-Clínica de Medicamentos/métodos , Vetores Genéticos/genética , Antivirais/farmacologia , Suramina/farmacologia , Efrina-B2/metabolismo , Efrina-B2/genética , Infecções por Henipavirus/virologia , Células Sf9 , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/metabolismo , Internalização do Vírus/efeitos dos fármacosRESUMO
Emerging highly pathogenic viruses can pose profound impacts on global health, the economy, and society. To meet that challenge, the National Institute of Allergy and Infectious Diseases (NIAID) established nine Antiviral Drug Discovery (AViDD) centers for early-stage identification and validation of novel antiviral drug candidates against viruses with pandemic potential. As part of this initiative, we established paired entry assays that simultaneously screen for inhibitors specifically targeting SARS-CoV-2 (SARS2), Lassa virus (LASV) and Machupo virus (MACV) entry. To do so we employed a dual pseudotyped virus (PV) infection system allowing us to screen â¼650,000 compounds efficiently and cost-effectively. Adaptation of these paired assays into 1536 well-plate format for ultra-high throughput screening (uHTS) resulted in the largest screening ever conducted in our facility, with over 2.4 million wells completed. The paired infection system allowed us to detect two PV infections simultaneously: LASV + MACV, MACV + SARS2, and SARS2 + LASV. Each PV contains a different luciferase reporter gene which enabled us to measure the infection of each PV exclusively, albeit in the same well. Each PV was screened at least twice utilizing different reporters, which allowed us to select the inhibitors specific to a particular PV and to exclude those that hit off targets, including cellular components or the reporter proteins. All assays were robust with an average Z' value ranging from 0.5 to 0.8. The primary screening of â¼650,000 compounds resulted in 1812, 1506, and 2586 unique hits for LASV, MACV, and SARS2, respectively. The confirmation screening narrowed this list further to 60, 40, and 90 compounds that are unique to LASV, MACV, and SARS2, respectively. Of these compounds, 8, 35, and 50 compounds showed IC50 value < 10 µM, some of which have much greater potency and excellent antiviral activity profiles specific to LASV, MACV, and SARS2, and none are cytotoxic. These selected compounds are currently being studied for their mechanism of action and to improve their specificity and potency through chemical modification.
Assuntos
Antivirais , Ensaios de Triagem em Larga Escala , Vírus Lassa , SARS-CoV-2 , Internalização do Vírus , Humanos , SARS-CoV-2/efeitos dos fármacos , Antivirais/farmacologia , Internalização do Vírus/efeitos dos fármacos , Vírus Lassa/efeitos dos fármacos , Ensaios de Triagem em Larga Escala/métodos , COVID-19/virologia , Descoberta de Drogas/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Tratamento Farmacológico da COVID-19 , Chlorocebus aethiops , AnimaisRESUMO
The chemokine co-receptors CXCR4 and CCR5 mediate HIV entry and signal transduction necessary for viral infection. However, to date only the CCR5 antagonist maraviroc is approved for treating HIV-1 infection. Given that approximately 50% of late-stage HIV patients also develop CXCR4-tropic virus, clinical anti-HIV CXCR4 antagonists are needed. Here, we describe a novel allosteric CXCR4 antagonist TIQ-15 which inhibits CXCR4-tropic HIV-1 infection of primary and transformed CD4 T cells. TIQ-15 blocks HIV entry with an IC50 of 13 nM. TIQ-15 also inhibits SDF-1α/CXCR4-mediated cAMP production, cofilin activation, and chemotactic signaling. In addition, TIQ-15 induces CXCR4 receptor internalization without affecting the levels of the CD4 receptor, suggesting that TIQ-15 may act through a novel allosteric site on CXCR4 for blocking HIV entry. Furthermore, TIQ-15 did not inhibit VSV-G pseudotyped HIV-1 infection, demonstrating its specificity in blocking CXCR4-tropic virus entry, but not CXCR4-independent endocytosis or post-entry steps. When tested against a panel of clinical isolates, TIQ-15 showed potent inhibition against CXCR4-tropic and dual-tropic viruses, and moderate inhibition against CCR5-tropic isolates. This observation was followed by a co-dosing study with maraviroc, and TIQ-15 demonstrated synergistic activity. In summary, here we describe a novel HIV-1 entry inhibitor, TIQ-15, which potently inhibits CXCR4-tropic viruses while possessing low-level synergistic activities against CCR5-tropic viruses. TIQ-15 could potentially be co-dosed with the CCR5 inhibitor maraviroc to block viruses of mixed tropisms.
Assuntos
Infecções por HIV , HIV-1 , Receptores CXCR4 , Internalização do Vírus , Humanos , Receptores CXCR4/antagonistas & inibidores , Receptores CXCR4/metabolismo , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Internalização do Vírus/efeitos dos fármacos , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Inibidores da Fusão de HIV/farmacologia , Maraviroc/farmacologia , Triazóis/farmacologia , Fármacos Anti-HIV/farmacologia , Células HEK293RESUMO
Many viruses contain surface spikes or protrusions that are essential for virus entry. These surface structures can thereby be targeted by antiviral drugs to treat viral infections. Nervous necrosis virus (NNV), a simple nonenveloped virus in the genus of betanodavirus, infects fish and damages aquaculture worldwide. NNV has 60 conspicuous surface protrusions, each comprising three protrusion domains (P-domain) of its capsid protein. NNV uses protrusions to bind to common receptors of sialic acids on the host cell surface to initiate its entry via the endocytic pathway. However, structural alterations of NNV in response to acidic conditions encountered during this pathway remain unknown, while detailed interactions of protrusions with receptors are unclear. Here, we used cryo-EM to discover that Grouper NNV protrusions undergo low-pH-induced compaction and resting. NMR and molecular dynamics (MD) simulations were employed to probe the atomic details. A solution structure of the P-domain at pH 7.0 revealed a long flexible loop (amino acids 311-330) and a pocket outlined by this loop. Molecular docking analysis showed that the N-terminal moiety of sialic acid inserted into this pocket to interact with conserved residues inside. MD simulations demonstrated that part of this loop converted to a ß-strand under acidic conditions, allowing for P-domain trimerization and compaction. Additionally, a low-pH-favored conformation is attained for the linker connecting the P-domain to the NNV shell, conferring resting protrusions. Our findings uncover novel pH-dependent conformational switching mechanisms underlying NNV protrusion dynamics potentially utilized for facilitating NNV entry, providing new structural insights into complex NNV-host interactions with the identification of putative druggable hotspots on the protrusion.
Assuntos
Proteínas do Capsídeo , Microscopia Crioeletrônica , Simulação de Dinâmica Molecular , Nodaviridae , Internalização do Vírus , Nodaviridae/efeitos dos fármacos , Nodaviridae/fisiologia , Nodaviridae/química , Concentração de Íons de Hidrogênio , Proteínas do Capsídeo/química , Proteínas do Capsídeo/metabolismo , Animais , Internalização do Vírus/efeitos dos fármacos , Antivirais/farmacologia , Antivirais/química , Doenças dos Peixes/virologia , Infecções por Vírus de RNA/virologiaRESUMO
Infection with respiratory syncytial virus (RSV) is a major cause of acute lower respiratory tract disease in young children and older people. Despite intensive efforts over the past few decades, no direct-acting small-molecule agents against RSV are available. Most small-molecule candidates targeting the RSV fusion (F) protein pose a considerable risk of inducing drug-resistant mutations. Here, we explored the in vitro and in vivo virological properties of the K394R variant, a cross-resistant mutant capable of evading multiple RSV fusion inhibitors. Our results demonstrated that the K394R variant is highly fusogenic in vitro and more pathogenic than the parental strain in vivo. The small molecule (2E,2'E)-N,N'-((1R,2S,3S)-3-hydroxycyclohexane-1,2-diyl)bis(3-(2-bromo-4-fluorophenyl) acrylamide) (CL-A3-7), a structurally optimized compound derived from a natural caffeoylquinic acid derivative, substantially reduced in vitro and in vivo infections of both wild-type RSV and the K394R variant. Mechanistically, CL-A3-7 significantly inhibited virus-cell fusion during RSV entry by blocking the interaction between the viral F protein and the cellular insulin-like growth factor 1 receptor (IGF1R). Collectively, these results indicate severe disease risks caused by the K394R variant and reveal a new anti-RSV mechanism to overcome K394R-associated resistance. IMPORTANCE: Respiratory syncytial virus (RSV) infection is a major public health concern, and many small-molecule candidates targeting the viral fusion (F) protein are associated with a considerable risk of inducing drug-resistant mutations. This study investigated virological features of the K394R variant, a mutant strain conferring resistance to multiple RSV fusion inhibitors. Our results demonstrated that the K394R variant is highly fusogenic in cell cultures and more pathogenic than the parental strain in mice. The small-molecule inhibitor CL-A3-7 substantially reduced in vitro and in vivo infections of both wild-type RSV and the K394R variant by blocking the interaction of viral F protein with its cellular receptor, showing a new mechanism of action for small-molecules to inhibit RSV infection and overcome K394R-associated resistance.
Assuntos
Antivirais , Farmacorresistência Viral , Infecções por Vírus Respiratório Sincicial , Vírus Sincicial Respiratório Humano , Proteínas Virais de Fusão , Internalização do Vírus , Internalização do Vírus/efeitos dos fármacos , Animais , Humanos , Antivirais/farmacologia , Farmacorresistência Viral/genética , Farmacorresistência Viral/efeitos dos fármacos , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/metabolismo , Proteínas Virais de Fusão/antagonistas & inibidores , Camundongos , Infecções por Vírus Respiratório Sincicial/tratamento farmacológico , Infecções por Vírus Respiratório Sincicial/virologia , Vírus Sincicial Respiratório Humano/efeitos dos fármacos , Vírus Sincicial Respiratório Humano/genética , Vírus Sincicial Respiratório Humano/fisiologia , Camundongos Endogâmicos BALB C , Linhagem Celular , FemininoRESUMO
Despite advancements in antiretroviral therapy (ART) suppressing HIV-1 replication, existing antiviral drugs pose limitations, including lifelong medication, frequent administration, side effects and viral resistance, necessitating novel HIV-1 treatment approaches. CD4, pivotal for HIV-1 entry, poses challenges for drug development due to neutralization and cytotoxicity concerns. Nevertheless, Ibalizumab, the sole approved CD4-specific antibody for HIV-1 treatment, reignites interest in exploring alternative anti-HIV targets, emphasizing CD4's potential value for effective drug development. Here, we explore anti-CD4 nanobodies, particularly Nb457 from a CD4-immunized alpaca. Nb457 displays high potency and broad-spectrum activity against HIV-1, surpassing Ibalizumab's efficacy. Strikingly, engineered trimeric Nb457 nanobodies achieve complete inhibition against live HIV-1, outperforming Ibalizumab and parental Nb457. Structural analysis unveils Nb457-induced CD4 conformational changes impeding viral entry. Notably, Nb457 demonstrates therapeutic efficacy in humanized female mouse models. Our findings highlight anti-CD4 nanobodies as promising HIV-1 therapeutics, with potential implications for advancing clinical treatment against this global health challenge.
Assuntos
Antígenos CD4 , Camelídeos Americanos , Anticorpos Anti-HIV , Infecções por HIV , HIV-1 , Anticorpos de Domínio Único , HIV-1/imunologia , HIV-1/efeitos dos fármacos , Anticorpos de Domínio Único/farmacologia , Anticorpos de Domínio Único/imunologia , Animais , Antígenos CD4/imunologia , Antígenos CD4/metabolismo , Humanos , Infecções por HIV/imunologia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Camelídeos Americanos/imunologia , Anticorpos Anti-HIV/imunologia , Anticorpos Anti-HIV/farmacologia , Camundongos , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Conformação Proteica , Feminino , Internalização do Vírus/efeitos dos fármacos , Células HEK293 , Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/uso terapêutico , Anticorpos MonoclonaisRESUMO
Since the largest and most fatal Ebola virus epidemic during 2014-2016, there have been several consecutive filoviral outbreaks in recent years, including those in 2021, 2022, and 2023. Ongoing outbreak prevalence and limited FDA-approved filoviral therapeutics emphasize the need for novel small molecule treatments. Here, we showcase the structure-activity relationship development of N-substituted pyrrole-based heterocycles and their potent, submicromolar entry inhibition against diverse filoviruses in a target-based pseudovirus assay. Inhibitor antiviral activity was validated using replication-competent Ebola, Sudan, and Marburg viruses. Mutational analysis was used to map the targeted region within the Ebola virus glycoprotein. Antiviral counter-screen and phospholipidosis assays were performed to demonstrate the reduced off-target activity of these filoviral entry inhibitors. Favorable antiviral potency, selectivity, and drug-like properties of the N-substituted pyrrole-based heterocycles support their potential as broad-spectrum antifiloviral treatments.