RESUMO
Virus proliferation involves gene replication inside infected cells and transmission to new target cells. Once positive-strand RNA virus has infected a cell, the viral genome serves as a template for copying ("stay-strategy") or is packaged into a progeny virion that will be released extracellularly ("leave-strategy"). The balance between genome replication and virion release determines virus production and transmission efficacy. The ensuing trade-off has not yet been well characterized. In this study, we use hepatitis C virus (HCV) as a model system to study the balance of the two strategies. Combining viral infection cell culture assays with mathematical modeling, we characterize the dynamics of two different HCV strains (JFH-1, a clinical isolate, and Jc1-n, a laboratory strain), which have different viral release characteristics. We found that 0.63% and 1.70% of JFH-1 and Jc1-n intracellular viral RNAs, respectively, are used for producing and releasing progeny virions. Analysis of the Malthusian parameter of the HCV genome (i.e., initial proliferation rate) and the number of de novo infections (i.e., initial transmissibility) suggests that the leave-strategy provides a higher level of initial transmission for Jc1-n, whereas, in contrast, the stay-strategy provides a higher initial proliferation rate for JFH-1. Thus, theoretical-experimental analysis of viral dynamics enables us to better understand the proliferation strategies of viruses, which contributes to the efficient control of virus transmission. Ours is the first study to analyze the stay-leave trade-off during the viral life cycle and the significance of the replication-release switching mechanism for viral proliferation.
Assuntos
Genoma Viral , Hepacivirus/genética , Interações Hospedeiro-Patógeno/genética , Envelhecimento/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Hepatite C , Humanos , Modelos Biológicos , Replicação Viral/genéticaRESUMO
Juglorubin is a natural dye isolated from the culture of Streptomyces sp. 3094, 815, and GW4184. It has been previously synthesized via the biomimetic dimerization of juglomycin C, a plausible genetic precursor. In this study, the derivatives of juglorubin, 1-O-acetyljuglorubin dimethyl ester and juglorubin dimethyl ester, were found to exhibit antiviral activity against hepatitis C virus (HCV) without exerting any remarkable cytotoxicity against host Huh-7 cells. They also inhibited liver X receptor α activation and lipid droplet accumulation in Huh-7 cells. These findings suggest that 1-O-acetyljuglorubin dimethyl ester and juglorubin dimethyl ester targeted the host factors required for HCV production.
Assuntos
Hepacivirus , Hepatite C , Humanos , Hepacivirus/genética , Linhagem Celular , Ésteres , Replicação Viral , Antivirais/farmacologiaRESUMO
Although aryl hydrocarbon receptors (AhRs) are related to the metabolic pathway of xenobiotics, recent studies have revealed that this receptor is also associated with the life cycle of viruses and inflammatory reactions. For example, flutamide, used to treat prostate cancer, inhibits hepatitis C virus proliferation by acting as an AhR antagonist, and methylated-pelargonidin, an AhR agonist, suppresses pro-inflammatory cytokine production. To discover a novel class of AhR ligands, we screened 1000 compounds derived from fungal metabolites using a reporter assay and identified methylsulochrin as a partial agonist of the aryl hydrocarbon receptor. Methylsulochrin was found to inhibit the production of hepatitis C virus (HCV) in Huh-7.5.1 cells. Methylsulochrin also suppressed the production of interleukin-6 in RAW264.7 cells. Furthermore, a preliminary structure-activity relationship study using sulochrin derivatives was performed. Our findings suggest the use of methylsulochrin derivatives as anti-HCV compounds with anti-inflammatory activity.
Assuntos
Antivirais , Receptores de Hidrocarboneto Arílico , Masculino , Humanos , Receptores de Hidrocarboneto Arílico/agonistas , Receptores de Hidrocarboneto Arílico/metabolismo , Antivirais/farmacologia , Flutamida/farmacologia , Anti-Inflamatórios/farmacologia , LigantesRESUMO
We have previously reported that neoechinulin B (1a), a prenylated indole diketopiperazine alkaloid, shows antiviral activities against hepatitis C virus (HCV) via the inactivation of the liver X receptors (LXRs) and the resultant disruption of double-membrane vesicles. In this study, a two-step synthesis of the diketopiperazine scaffold of 1a was achieved by the base-induced coupling of 1,4-diacetyl-3-{[(tert-butyldimethylsilyl)oxy]methyl}piperazine-2,5-dione with aldehydes, followed by the treatment of the resultant coupling products with tetra-n-butylammonium fluoride. Compound 1a and its 16 derivatives 1b-q were prepared using this method. Furthermore, variecolorin H, a related alkaloid, was obtained by the acid treatment of 1a in MeOH. The antiviral evaluation of 1a and its derivatives revealed that 1a, 1c, 1d, 1h, 1j, 1l, and 1o exhibited both anti-HCV and anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) activities. The results of this study indicate that the exomethylene moiety on the diketopiperazine ring is important for the antiviral activities. The antiviral compounds can inhibit the production of HCV and SARS-CoV-2 by inactivating LXRs.
Assuntos
Alcaloides/farmacologia , Antivirais/farmacologia , Hepacivirus/efeitos dos fármacos , Piperazinas/farmacologia , SARS-CoV-2/efeitos dos fármacos , Alcaloides/síntese química , Alcaloides/química , Antivirais/síntese química , Antivirais/química , Linhagem Celular Tumoral , Dicetopiperazinas/química , Dicetopiperazinas/farmacologia , Humanos , Receptores X do Fígado/antagonistas & inibidores , Estrutura Molecular , Piperazinas/síntese química , Piperazinas/química , Relação Estrutura-Atividade , Transcrição Gênica/efeitos dos fármacosRESUMO
The liver X receptor is a nuclear hormone receptor that regulates lipid metabolism. Previously, we had demonstrated the antiviral properties of a liver X receptor antagonist associated with the hepatitis C virus and severe acute respiratory syndrome coronavirus 2. In this study, we screened a chemical library and identified two potential liver X receptor antagonists. Spectroscopic analysis revealed that the structures of both antagonists (compounds 1 and 2) were cyclic dimer and trimer of esters, respectively, that consisted of phthalate and 1,6-hexane diol. This study is the first to report the structure of the cyclic trimer of phthalate ester. Further experiments revealed that the compounds were impurities of solvents used for purification, although their source could not be traced. Both phthalate esters exhibited anti-hepatitis C virus activity, whereas the cyclic dimer showed anti-severe acute respiratory syndrome coronavirus 2 activity. Cyclic phthalate derivatives may constitute a novel class of liver X receptor antagonists and broad-spectrum antivirals.
Assuntos
COVID-19 , Ésteres , Antivirais/farmacologia , Ésteres/farmacologia , Hepacivirus , Hexanos , Humanos , Receptores X do Fígado , Ácidos Ftálicos , Receptores Citoplasmáticos e Nucleares , SARS-CoV-2 , SolventesRESUMO
Sodium taurocholate cotransporting polypeptide (NTCP) is expressed at the surface of human hepatocytes and functions as an entry receptor of hepatitis B virus (HBV). Recently, we have reported that epidermal growth factor receptor (EGFR) is involved in NTCP-mediated viral internalization during the cell entry process. Here, we analyzed which function of EGFR is essential for mediating HBV internalization. In contrast to the reported crucial function of EGFR-downstream signaling for the entry of hepatitis C virus (HCV), blockade of EGFR-downstream signaling proteins, including mitogen-activated protein kinase (MAPK), phosphoinositide 3-kinase (PI3K), and signal transducer and activator of transcription (STAT), had no or only minor effects on HBV infection. Instead, deficiency of EGFR endocytosis resulting from either a deleterious mutation in EGFR or genetic knockdown of endocytosis adaptor molecules abrogated internalization of HBV via NTCP and prevented viral infection. EGFR activation triggered a time-dependent relocalization of HBV preS1 to the early and late endosomes and to lysosomes in concert with EGFR transport. Suppression of EGFR ubiquitination by site-directed mutagenesis or by knocking down two EGFR-sorting molecules, signal-transducing adaptor molecule (STAM) and lysosomal protein transmembrane 4ß (LAPTM4B), suggested that EGFR transport to the late endosome is critical for efficient HBV infection. Cumulatively, these results support the idea that the EGFR endocytosis/sorting machinery drives the translocation of NTCP-bound HBV from the cell surface to the endosomal network, which eventually enables productive viral infection.
Assuntos
Endocitose/genética , Endossomos/genética , Receptores ErbB/genética , Hepatite B/genética , Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/genética , Complexos Endossomais de Distribuição Requeridos para Transporte/química , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Endossomos/química , Receptores ErbB/química , Células Hep G2 , Hepacivirus/química , Hepacivirus/genética , Hepacivirus/patogenicidade , Hepatite B/metabolismo , Hepatite B/virologia , Vírus da Hepatite B/química , Vírus da Hepatite B/genética , Vírus da Hepatite B/patogenicidade , Hepatócitos/metabolismo , Hepatócitos/virologia , Humanos , MAP Quinase Quinase 1/genética , Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas Oncogênicas/química , Proteínas Oncogênicas/genética , Transportadores de Ânions Orgânicos Dependentes de Sódio , Fosfatidilinositol 3-Quinases/genética , Fosfoproteínas/química , Fosfoproteínas/genética , Fatores de Transcrição STAT/genética , Simportadores , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/genética , Internalização do VírusRESUMO
4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) and 4'-ethynyl-2'-deoxyadenosine (EdA) are nucleoside analogues which inhibit human immunodeficiency virus type 1 (HIV-1) reverse transcriptase. EdAP, a cyclosaligenyl (cycloSal) phosphate derivative of EdA, inhibits the replication of the influenza A virus. The common structural feature of these compounds is the ethynyl group at the 4'-position. In this study, these nucleoside analogues were prepared by a common synthetic strategy starting from the known 1,2-di-O-acetyl-D-ribofuranose. Biological evaluation of EdAP revealed that this compound reduced hepatitis B virus (HBV) replication dose-dependently without cytotoxicity against host cells tested in this study.
Assuntos
Antivirais/síntese química , Nucleotídeos de Desoxiadenina/síntese química , Desoxiadenosinas/síntese química , Vírus da Hepatite B/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Antivirais/farmacologia , Linhagem Celular , Nucleotídeos de Desoxiadenina/farmacologia , Desoxiadenosinas/farmacologia , Vírus da Hepatite B/fisiologia , HumanosRESUMO
Viruses hijack and modify host cell functions to maximize viral proliferation. Hepatitis C virus (HCV) reorganizes host cell metabolism to produce specialized membrane structures and to modify organelles such as double-membrane vesicles and enlarged lipid droplets (LDs), thereby enabling virus replication and assembly. However, the molecular bases of these host-HCV interactions are largely unknown. Here, using a chemical screen, we demonstrate that the benzamide derivative flutamide reduces the host capacity to produce infectious HCV. Flutamide disrupted the formation of enlarged LDs in HCV-infected cells, thereby abolishing HCV assembly. We also report that aryl hydrocarbon receptor (AhR), a known flutamide target, plays a key role in mediating LD accumulation and HCV production. This AhR function in lipid production was also observed in HCV-uninfected Huh-7 cells and primary human hepatocytes, suggesting that AhR signaling regulates lipid accumulation independently of HCV infection. We further observed that a downstream activity, that of cytochrome P450 1A1 (CYP1A1), was the primary regulator of AhR-mediated lipid production. Specifically, blockade of AhR-induced CYP1A1 up-regulation counteracted LD overproduction, and overproduction of CYP1A1, but not of CYP1B1, in AhR-inactivated cells restored lipid accumulation. Of note, HCV infection up-regulated the AhR-CYP1A1 pathway, resulting in the accumulation of enlarged LDs. In conclusion, we demonstrate that the AhR-CYP1A1 pathway has a significant role in lipid accumulation, a hallmark of HCV infection that maximizes progeny virus production. Our chemical-genetic analysis reveals a new strategy and lead compounds to control hepatic lipid accumulation as well as HCV infection.
Assuntos
Citocromo P-450 CYP1A1/metabolismo , Hepacivirus/fisiologia , Metabolismo dos Lipídeos , Receptores de Hidrocarboneto Arílico/metabolismo , Montagem de Vírus , Linhagem Celular , Flutamida/farmacologia , Hepacivirus/efeitos dos fármacos , Humanos , Gotículas Lipídicas/efeitos dos fármacos , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Ligação Proteica , Montagem de Vírus/efeitos dos fármacosRESUMO
Antiviral treatments targeting the coronavirus disease 2019 are urgently required. We screened a panel of already approved drugs in a cell culture model of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and identified two new agents having higher antiviral potentials than the drug candidates such as remdesivir and chroloquine in VeroE6/TMPRSS2 cells: the anti-inflammatory drug cepharanthine and human immunodeficiency virus protease inhibitor nelfinavir. Cepharanthine inhibited SARS-CoV-2 entry through the blocking of viral binding to target cells, while nelfinavir suppressed viral replication partly by protease inhibition. Consistent with their different modes of action, synergistic effect of this combined treatment to limit SARS-CoV-2 proliferation was highlighted. Mathematical modeling in vitro antiviral activity coupled with the calculated total drug concentrations in the lung predicts that nelfinavir will shorten the period until viral clearance by 4.9 days and the combining cepharanthine/nelfinavir enhanced their predicted efficacy. These results warrant further evaluation of the potential anti-SARS-CoV-2 activity of cepharanthine and nelfinavir.