Learning a lesson from a famous puppet player Susumu Tange : A good practice of "Practical training of child-care skills" in Kyoto Women's college

textabstractCoronaviruses can cause respiratory and enteric disease in a wide variety of human and animal hosts. The 2003 outbreak of severe acute respiratory syndrome (SARS) first demonstrated the potentially lethal consequences of zoonotic coronavirus infections in humans. In 2012, a similar previously unknown coronavirus emerged, Middle East respiratory syndrome coronavirus (MERS-CoV), thus far causing over 650 laboratory-confirmed infections, with an unexplained steep rise in the number of cases being recorded over recent months. The human MERS fatality rate of∼30% is alarmingly high, even though many deaths were associated with underlying medical conditions. Registered therapeutics for the treatment of coronavirus infections are not available. Moreover, the pace of drug development and registration for human use is generally incompatible with strategies to combat emerging infectious diseases. Therefore, we have screened a library of 348 FDA-approved drugs for anti-MERS-CoV activity in cell culture. If such compounds proved sufficiently potent, their efficacy might be directly assessed in MERS patients. We identified four compounds (chloroquine, chlorpromazine, loperamide, and lopinavir) inhibiting MERS-CoV replication in the lowmicromolar range (50% effective concentrations [EC50s], 3 to 8 μM). Moreover, these compounds also inhibit the replication of SARS coronavirus and human coronavirus 229E. Although their protective activity (alone or in combination) remains to be assessed in animal models, our findings may offer a starting point for treatment of patients infected with zoonotic coronaviruses like MERS-CoV. Although they may not necessarily reduce viral replication to very low levels, a moderate viral load reduction may create a window during which to mount a protective immune response. Copyrigh

Design, synthesis and evaluation of a series of acyclic fleximer nucleoside analogues with anti-coronavirus activity

A series of doubly flexible nucleoside analogues were designed based on the acyclic sugar scaffold of acyclovir and the flex-base moiety found in the fleximers. The target compounds were evaluated for their antiviral potential and found to inhibit several coronaviruses. Significantly, compound 2 displayed selective antiviral activity (CC50 >3× EC50) towards human coronavirus (HCoV)-NL63 and Middle East respiratory syndrome-coronavirus, but not severe acute respiratory syndrome-coronavirus. In the case of HCoV-NL63 the activity was highly promising with an EC50 100μM. As such, these doubly flexible nucleoside analogues are viewed as a novel new class of drug candidates with potential for potent inhibition of coronaviruses.publisher: Elsevier articletitle: Design, synthesis and evaluation of a series of acyclic fleximer nucleoside analogues with anti-coronavirus activity journaltitle: Bioorganic & Medicinal Chemistry Letters articlelink: http://dx.doi.org/10.1016/j.bmcl.2015.05.039 content_type: article copyright: Copyright © 2015 Elsevier Ltd. All rights reserved.status: publishe

A unifying structural and functional model of the coronavirus replication organelle: Tracking down RNA synthesis

Zoonotic coronavirus (CoV) infections, such as those responsible for the current severe acute respiratory syndrome-CoV 2 (SARS-CoV-2) pandemic, cause grave international public health concern. In infected cells, the CoV RNA-synthesizing machinery associates with modified endoplasmic reticulum membranes that are transformed into the viral replication organelle (RO). Although double-membrane vesicles (DMVs) appear to be a pan-CoV RO element, studies to date describe an assortment of additional CoV-induced membrane structures. Despite much speculation, it remains unclear which RO element(s) accommodate viral RNA synthesis. Here we provide detailed 2D and 3D analyses of CoV ROs and show that diverse CoVs essentially induce the same membrane modifications, including the small open double-membrane spherules (DMSs) previously thought to be restricted to gamma- and delta-CoV infections and proposed as sites of replication. Metabolic labeling of newly synthesized viral RNA followed by quantitative electron microscopy (EM) autoradiography revealed abundant viral RNA synthesis associated with DMVs in cells infected with the beta-CoVs Middle East respiratory syndrome-CoV (MERS-CoV) and SARS-CoV and the gamma-CoV infectious bronchitis virus. RNA synthesis could not be linked to DMSs or any other cellular or virus-induced structure. Our results provide a unifying model of the CoV RO and clearly establish DMVs as the central hub for viral RNA synthesis and a potential drug target in CoV infection

Design, synthesis and evaluation of a series of acyclic fleximer nucleoside analogues with anti-coronavirus activity

A series of doubly flexible nucleoside analogues were designed based on the acyclic sugar scaffold of acyclovir and the flex-base moiety found in the fleximers. The target compounds were evaluated for their antiviral potential and found to inhibit several coronaviruses. Significantly, compound 2 displayed selective antiviral activity (CC(50) > 3x EC(50)) towards human coronavirus (HCoV)-NL63 and Middle East respiratory syndrome-coronavirus, but not severe acute respiratory syndrome-coronavirus. In the case of HCoV-NL63 the activity was highly promising with an EC(50) < 10 μM and a CC(50) > 100 μM. As such, these doubly flexible nucleoside analogues are viewed as a novel new class of drug candidates with potential for potent inhibition of coronaviruses

Cyclosporin A inhibits the replication of diverse coronaviruses

Low micromolar, non-cytotoxic concentrations of cyclosporin A (CsA) strongly affected the replication of severe acute respiratory syndrome coronavirus (SARS-CoV), human coronavirus 229E and mouse hepatitis virus in cell culture, as was evident from the strong inhibition of GFP reporter gene expression and a reduction of up to 4 logs in progeny titres. Upon high-multiplicity infection, CsA treatment rendered SARS-CoV RNA and protein synthesis almost undetectable, suggesting an early block in replication. siRNA-mediated knockdown of the expression of the prominent CsA targets cyclophilin A and B did not affect SARS-CoV replication, suggesting either that these specific cyclophilin family members are dispensable or that the reduced expression levels suffice to support replication

Screening of an FDA-approved compound library identifies four small-molecule Inhibitors of Middle East respiratory syndrome coronavirus replication in cell culture

Coronaviruses can cause respiratory and enteric disease in a wide variety of human and animal hosts. The 2003 outbreak of severe acute respiratory syndrome (SARS) first demonstrated the potentially lethal consequences of zoonotic coronavirus infections in humans. In 2012, a similar previously unknown coronavirus emerged, Middle East respiratory syndrome coronavirus (MERS-CoV), thus far causing over 650 laboratory-confirmed infections, with an unexplained steep rise in the number of cases being recorded over recent months. The human MERS fatality rate of ∼30% is alarmingly high, even though many deaths were associated with underlying medical conditions. Registered therapeutics for the treatment of coronavirus infections are not available. Moreover, the pace of drug development and registration for human use is generally incompatible with strategies to combat emerging infectious diseases. Therefore, we have screened a library of 348 FDA-approved drugs for anti-MERS-CoV activity in cell culture. If such compounds proved sufficiently potent, their efficacy might be directly assessed in MERS patients. We identified four compounds (chloroquine, chlorpromazine, loperamide, and lopinavir) inhibiting MERS-CoV replication in the low-micromolar range (50% effective concentrations [EC50s], 3 to 8 μM). Moreover, these compounds also inhibit the replication of SARS coronavirus and human coronavirus 229E. Although their protective activity (alone or in combination) remains to be assessed in animal models, our findings may offer a starting point for treatment of patients infected with zoonotic coronaviruses like MERS-CoV. Although they may not necessarily reduce viral replication to very low levels, a moderate viral load reduction may create a window during which to mount a protective immune response.status: publishe