An analogue of the antibiotic teicoplanin prevents flavivirus entry in vitro.

There is an urgent need for potent inhibitors of dengue virus (DENV) replication for the treatment and/or prophylaxis of infections with this virus. We here report on an aglycon analogue of the antibiotic teicoplanin (code name LCTA-949) that inhibits DENV-induced cytopathic effect (CPE) in a dose-dependent manner. Virus infection was completely inhibited at concentrations that had no adverse effect on the host cells. These findings were corroborated by quantification of viral RNA levels in culture supernatant. Antiviral activity was also observed against other flaviviruses such as the yellow fever virus and the tick-borne encephalitis virus (TBEV). In particular, potent antiviral activity was observed against TBEV. Time-of-drug-addition experiments indicated that LCTA-949 inhibits an early stage in the DENV replication cycle; however, a virucidal effect was excluded. This observation was corroborated by the fact that LCTA-949 lacks activity on DENV subgenomic replicon (that does not encode structural proteins) replication. Using a microsopy-based binding and fusion assay employing DiD-labeled viruses, it was shown that LCTA-949 targets the early stage (binding/entry) of the infection. Moreover, LCTA-949 efficiently inhibits infectivity of DENV particles pre-opsonized with antibodies, thus potentially also inhibiting antibody-dependent enhancement (ADE). In conclusion, LCTA-949 exerts in vitro activity against several flaviviruses and does so (as shown for DENV) by interfering with an early step in the viral replication cycle.Fil: De Burghgraeve, Tine. Katholikie Universiteit Leuven; BélgicaFil: Kaptein, Suzanne J. F.. Katholikie Universiteit Leuven; BélgicaFil: Ayala Nunez, Nilda V.. University of Groningen; Países BajosFil: Mondotte, Juan Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Pastorino, Boris. Université de la Méditerranée; FranciaFil: Printsevskaya, Svetlana S.. Russian Academy of Medical Sciences; RusiaFil: de Lamballerie, Xavier. Université de la Méditerranée; FranciaFil: Jacobs, Michael. Royal Free & University College Medical School; Reino UnidoFil: Preobrazhenskaya, Maria. Russian Academy of Medical Sciences; RusiaFil: Gamarnik, Andrea Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Smit, Jolanda M.. University of Groningen; Países BajosFil: Neyts, Johan. Katholikie Universiteit Leuven; Bélgic

Broad Antiviral Activity of Carbohydrate-Binding Agents against the Four Serotypes of Dengue Virus in Monocyte-Derived Dendritic Cells

BACKGROUND: Dendritic cells (DC), present in the skin, are the first target cells of dengue virus (DENV). Dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (DC-SIGN) is present on DC and recognizes N-glycosylation sites on the E-glycoprotein of DENV. Thus, the DC-SIGN/E-glycoprotein interaction can be considered as an important target for inhibitors of viral replication. We evaluated various carbohydrate-binding agents (CBAs) against all four described serotypes of DENV replication in Raji/DC-SIGN(+) cells and in monocyte-derived DC (MDDC). METHODOLOGY/PRINCIPAL FINDINGS: A dose-dependent anti-DENV activity of the CBAs Hippeastrum hybrid (HHA), Galanthus nivalis (GNA) and Urtica dioica (UDA), but not actinohivin (AH) was observed against all four DENV serotypes as analyzed by flow cytometry making use of anti-DENV antibodies. Remarkably, the potency of the CBAs against DENV in MDDC cultures was significantly higher (up to 100-fold) than in Raji/DC-SIGN(+) cells. Pradimicin-S (PRM-S), a small-size non-peptidic CBA, exerted antiviral activity in MDDC but not in Raji/DC-SIGN(+) cells. The CBAs act at an early step of DENV infection as they bind to the viral envelope of DENV and subsequently prevent virus attachment. Only weak antiviral activity of the CBAs was detected when administered after the virus attachment step. The CBAs were also able to completely prevent the cellular activation and differentiation process of MDDC induced upon DENV infection. CONCLUSIONS/SIGNIFICANCE: The CBAs exerted broad spectrum antiviral activity against the four DENV serotypes, laboratory-adapted viruses and low passage clinical isolates, evaluated in Raji/DC-SIGN(+) cells and in primary MDDC

Animal models for COVID-19

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the aetiological agent of coronavirus disease 2019 (COVID-19), an emerging respiratory infection caused by the introduction of a novel coronavirus into humans late in 2019 (frst detected in Hubei province, China). As of 18 September 2020, SARS-CoV-2 has spread to 215 countries, has infected more than 30 million people and has caused more than 950,000 deaths. As humans do not have pre-existing immunity to SARS-CoV-2, there is an urgent need to develop therapeutic agents and vaccines to mitigate the current pandemic and to prevent the re-emergence of COVID-19. In February 2020, the World Health Organization (WHO) assembled an international panel to develop animal models for COVID-19 to accelerate the testing of vaccines and therapeutic agents. Here we summarize the fndings to date and provides relevant information for preclinical testing of vaccine candidates and therapeutic agents for COVID-19.info:eu-repo/semantics/acceptedVersio

STAT2 signaling restricts viral dissemination but drives severe pneumonia in SARS-CoV-2 infected hamsters

Emergence of SARS-CoV-2 causing COVID-19 has resulted in hundreds of thousands of deaths. In search for key targets of effective therapeutics, robust animal models mimicking COVID-19 in humans are urgently needed. Here, we show that Syrian hamsters, in contrast to mice, are highly permissive to SARS-CoV-2 and develop bronchopneumonia and strong inflammatory responses in the lungs with neutrophil infiltration and edema, further confirmed as consolidations visualized by micro-CT alike in clinical practice. Moreover, we identify an exuberant innate immune response as key player in pathogenesis, in which STAT2 signaling plays a dual role, driving severe lung injury on the one hand, yet restricting systemic virus dissemination on the other. Our results reveal the importance of STAT2-dependent interferon responses in the pathogenesis and virus control during SARS-CoV-2 infection and may help rationalizing new strategies for the treatment of COVID-19 patients. SARS-CoV-2 infection can result in severe lung inflammation and pathology, but host response remains incompletely understood. Here the authors show in Syrian hamsters that STAT2 signaling restricts systemic virus dissemination but also drives severe lung injury, playing a dual role in SARS-CoV-2 infection

Scaffold Morphing Approach To Expand the Toolbox of Broad-Spectrum Antivirals Blocking Dengue/Zika Replication

We have recently discovered a family of 2,6-diaminopurine derivatives acting as DENV inhibitors by targeting an allosteric pocket on the thumb of the viral NS5 polymerase. Although the following target-based optimization allowed conversion of the hits into broad-spectrum DENV/ZIKV inhibitors, no improvement of the antiviral potency was reached. Herein, we applied a phenotypic scaffold-morphing approach to explore additional biologically relevant chemical space around the original hits by converting the flat purine derivatives into more complex chemotypes characterized by a higher degree of saturation. A new microwave-assisted one-pot three-step protocol was also developed to quickly generate chemotypes 6 and 7. Cell-based phenotypic screening allowed identification of promising antiflaviviral agents belonging to different chemotypes. Compound 9d emerged as the most promising broad-spectrum antiviral, being 6 times more potent than ribavirin (RBV) against DENV and 3 times more potent than 7-deaza-2′-C-methyladenosine (7DMA) against ZIKV with good selectivity indexes (>46 and >41, respectively)

Human stem cell-derived hepatocyte-like cells support Zika virus replication and provide a relevant model to assess the efficacy of potential antivirals.

Zika virus (ZIKV) infection during pregnancy has been extensively linked to microcephaly in newborns. High levels of ZIKV RNA were, however, also detected in mice and non-human primates in organs other than the brain, such as the liver. As ZIKV is a flavivirus closely related to the dengue and yellow fever virus, which are known to cause hepatitis, we here examined whether human hepatocytes are susceptible to ZIKV infection. We demonstrated that both human pluripotent stem cell (hPSC)-derived hepatocyte-like cells (HLCs) and the Huh7 hepatoma cell line support the complete ZIKV replication cycle. Of three antiviral molecules that inhibit ZIKV infection in Vero cells, only 7-deaza-2'-C-methyladenosine (7DMA) inhibited ZIKV replication in hPSC-HLCs, while all drugs inhibited ZIKV infection in Huh7 cells. ZIKV-infected hPSC-HLCs but not Huh7 cells mounted an innate immune and NFκβ response, which may explain the more extensive cytopathic effect observed in Huh7 cells. In conclusion, ZIKV productively infects human hepatocytes in vitro. However, significant differences in the innate immune response against ZIKV and antiviral drug sensitivity were observed when comparing hPSC-HLCs and hepatoma cells, highlighting the need to assess ZIKV infection as well as antiviral activity not only in hepatoma cells, but also in more physiologically relevant systems

Identification of Broad-Spectrum Dengue/Zika Virus Replication Inhibitors by Functionalization of Quinoline and 2,6-Diaminopurine Scaffolds

Social and demographic changes across the world over the past 50 years have resulted in significant outbreaks of arboviruses such as dengue virus (DENV) and Zika virus (ZIKV). Despite the increased threat of infection, no approved drugs or fully protective vaccines are available to counteract the spread of DENV and ZIKV. The development of "broad-spectrum" antivirals (BSAs) that target common components of multiple viruses can be a more effective strategy to limit the rapid emergence of viral pathogens than the classic "one-bug/one-drug" approach. Starting from previously identified multitarget DENV inhibitors, herein we report the identification of novel 2,6-diaminopurine derivatives that are able to block the replication of both Zika virus and all serotypes of dengue virus (DENV 1-4) in infected cells

The Rat Cytomegalovirus R33-Encoded G Protein-Coupled Receptor Signals in a Constitutive Fashion

The rat cytomegalovirus (RCMV) R33 gene is conserved among all betaherpesviruses and encodes a protein (pR33) that shows sequence similarity with chemokine-binding G protein-coupled receptors (GPCRs). Previously, the physiological significance of the R33 gene was demonstrated by the finding that an RCMV strain with R33 deleted is severely attenuated in vivo and is unable to either enter or replicate in the salivary glands of infected rats. Here, we report that RCMV pR33 is expressed as a functional GPCR that signals in an agonist-independent manner in both COS-7 and Rat2 cells. Transient expression of pR33 in COS-7 cells results in constitutive activation of phospholipase C (PLC) due to coupling to G proteins of the G(q) class. Interestingly, PLC activation is partially inhibited by cotransfection with G(α)-transducin subunits, which indicates the involvement of G(βγ) as well as Gα subunits in pR33-mediated signaling. Surprisingly, PLC activation is also partially inhibited by addition of pertussis toxin (PTX), suggesting that pR33 activates not only G(q) but also G(i/0) proteins. The constitutive activation of G(i/0) proteins by pR33 is further demonstrated by the PTX-sensitive decrease of CRE-mediated transcription and the PTX-sensitive increase of both NF-κB- and SRE-mediated transcription. In contrast to its homolog of human herpesvirus 6B (pU12), pR33 does not bind RANTES

Antiviral activity of HHA, GNA and UDA in MDDC and in Raji/DC-SIGN⁺ cells.

<p>MDDC (▾) or Raji/DC-SIGN⁺ cells (□) were infected with DENV-1, DENV-2, DENV-3 or DENV-4. Cells were incubated with increasing concentrations of HHA, GNA, UDA and viral infectivity was quantified by flow cytometry using 5 µg/ml anti-DENV mAb (clone 2H2). Data represent the percentage of inhibition of viral replication relative to the positive control (DENV-infected cells). Each value represents the mean ± SEM of 3 to 8 independent experiments.</p