The inhaled steroid ciclesonide blocks SARS-CoV-2 RNA replication by targeting viral replication-transcription complex in culture cells
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Abstract
We screened steroid compounds to obtain a drug expected to block host inflammatory responses and MERS-CoV replication. Ciclesonide, an inhaled corticosteroid, suppressed replication of MERS-CoV and other coronaviruses, including SARS-CoV-2, the cause of COVID-19, in cultured cells. The effective concentration (EC 90 ) of ciclesonide for SARS-CoV-2 in differentiated human bronchial tracheal epithelial cells was 0.55 μM. Ciclesonide inhibited formation of double membrane vesicles, which anchor the viral replication-transcription complex in cells. Eight consecutive passages of 43 SARS-CoV-2 isolates in the presence of ciclesonide generated 15 resistant mutants harboring single amino acid substitutions in non-structural protein 3 (nsp3) or nsp4. Of note, ciclesonide still suppressed replication of all these mutants by 90% or more, suggesting that these mutants cannot completely overcome ciclesonide blockade. These observations indicate that the suppressive effect of ciclesonide on viral replication is specific to coronaviruses, highlighting it as a candidate drug for the treatment of COVID-19 patients.
Importance
The outbreak of SARS-CoV-2, the cause of COVID-19, is ongoing. To identify the effective antiviral agents to combat the disease is urgently needed. In the present study, we found that an inhaled corticosteroid, ciclesonide suppresses replication of coronaviruses, including beta-coronaviruses (MHV-2, MERS-CoV, SARS-CoV, and SARS-CoV-2) and an alpha-coronavirus (HCoV-229E) in cultured cells. The inhaled ciclesonide is safe; indeed, it can be administered to infants at high concentrations. Thus, ciclesonide is expected to be a broad-spectrum antiviral drug that is effective against many members of the coronavirus family. It could be prescribed for the treatment of MERS, and COVID-19.
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SciScore for 10.1101/2020.08.22.258459: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources The cells were then incubated with a mixture of rabbit anti-SARS-nsp4 (1:500; ab181620; Abcam, USA) and mouse anti-dsRNA (1:1000; J2-1709; Scicons, Hungary) antibodies for 1 h at RT, washed three times with PBS, and incubated for 1 h at RT with a mixture of Alexa Fluor 594 conjugated anti-rabbit IgG (1:500; A11012; ThermoFisher, USA) and Alexa Fluor 488-conjugated anti-mouse IgG (1:500; A10680; ThermoFisher, USA). anti-SARS-nsp4suggested: Noneanti-dsRNAsugg…SciScore for 10.1101/2020.08.22.258459: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources The cells were then incubated with a mixture of rabbit anti-SARS-nsp4 (1:500; ab181620; Abcam, USA) and mouse anti-dsRNA (1:1000; J2-1709; Scicons, Hungary) antibodies for 1 h at RT, washed three times with PBS, and incubated for 1 h at RT with a mixture of Alexa Fluor 594 conjugated anti-rabbit IgG (1:500; A11012; ThermoFisher, USA) and Alexa Fluor 488-conjugated anti-mouse IgG (1:500; A10680; ThermoFisher, USA). anti-SARS-nsp4suggested: Noneanti-dsRNAsuggested: (Millipore Cat# MABE1134, RRID:AB_2819101)anti-rabbit IgGsuggested: (Molecular Probes Cat# A-11012, RRID:AB_141359)anti-mouse IgGsuggested: (Thermo Fisher Scientific Cat# A-10680, RRID:AB_2534062)Experimental Models: Cell Lines Sentences Resources Cells and viruses: Hep-2, HeLa229, MDCK, Calu-3, Vero, Vero/TMPRSS2 and VeroE6/TMPRSS2 cells were maintained in Dulbecco’s modified Eagle medium high glucose (DMEM, Sigma-Aldrich, USA), and DBT cells were maintained in DMEM (Nissui, Japan), supplemented with 5% fetal bovine serum (Gibco-BRL, USA). Hep-2suggested: NoneMDCKsuggested: CLS Cat# 602280/p823_MDCK_(NBL-2, RRID:CVCL_0422)Verosuggested: CLS Cat# 605372/p622_VERO, RRID:CVCL_0059)VeroE6/TMPRSS2suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)BHK-21 cells were grown in a single well of a six-well plate in 10% FCS-MEM and transfected with 3 μg of BAC plasmid with Lipofectamine 3,000 (Thermo Fisher, USA) BHK-21suggested: NoneThe supernatants were collected and propagated once using Vero/TMPRSS2 cells. Vero/TMPRSS2suggested: JCRB Cat# JCRB1818, RRID:CVCL_YQ48)Vero and VeroE6/TMPSS2 cells were used to passage MERS-CoV and SARS-CoV-2, respectively. VeroE6/TMPSS2suggested: NoneSoftware and Algorithms Sentences Resources Indexed libraries were then converted and sequenced (150-bp paired-end reads) using the DNBSEQ-G400 (MGI Tech., Shenzhen, China; operated by GENEWIZ, South Plainfield, NJ, USA). GENEWIZsuggested: (GENEWIZ, RRID:SCR_003177)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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