Drug synergy of combinatory treatment with remdesivir and the repurposed drugs fluoxetine and itraconazole effectively impairs SARS‐CoV‐2 infection in vitro
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Abstract
The SARS‐COV‐2 pandemic and the global spread of coronavirus disease 2019 (COVID‐19) urgently call for efficient and safe antiviral treatment strategies. A straightforward approach to speed up drug development at lower costs is drug repurposing. Here, we investigated the therapeutic potential of targeting the interface of SARS CoV‐2 with the host via repurposing of clinically licensed drugs and evaluated their use in combinatory treatments with virus‐ and host‐directed drugs in vitro.
Experimental Approach
We tested the antiviral potential of the antifungal itraconazole and the antidepressant fluoxetine on the production of infectious SARS‐CoV‐2 particles in the polarized Calu‐3 cell culture model and evaluated the added benefit of a combinatory use of these host‐directed drugs with the direct acting antiviral remdesivir, an inhibitor of viral RNA polymerase.
Key Results
Drug treatments were well‐tolerated and potently impaired viral replication. Importantly, both itraconazole–remdesivir and fluoxetine–remdesivir combinations inhibited the production of infectious SARS‐CoV‐2 particles > 90% and displayed synergistic effects, as determined in commonly used reference models for drug interaction.
Conclusion and Implications
Itraconazole–remdesivir and fluoxetine–remdesivir combinations are promising starting points for therapeutic options to control SARS‐CoV‐2 infection and severe progression of COVID‐19.
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SciScore for 10.1101/2020.10.16.342410: (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 Virus plaques were visualized by staining with neutral red, and virus titers were calculated as plaque-forming units (PFU) per mL. 2.5 Data and statistical analysis: A priori power analysis using G*Power 3.1 (Faul et al., 2007)) was performed to determine the sample sizes required to detect > 90% reduction in virus titers at powers > 0.8. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources 2.1 Cells and SARS-CoV-2 isolate: The human bronchial epithelial cell lines Calu-3 and the Vero E6 cells were … SciScore for 10.1101/2020.10.16.342410: (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 Virus plaques were visualized by staining with neutral red, and virus titers were calculated as plaque-forming units (PFU) per mL. 2.5 Data and statistical analysis: A priori power analysis using G*Power 3.1 (Faul et al., 2007)) was performed to determine the sample sizes required to detect > 90% reduction in virus titers at powers > 0.8. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources 2.1 Cells and SARS-CoV-2 isolate: The human bronchial epithelial cell lines Calu-3 and the Vero E6 cells were cultivated in Dulbecco’s modified Eagle’s medium (DMEM) with 10% standardized fetal bovine serum (FBS Superior; Merck), 2 mM L-glutamine, 100 U/mL penicillin, 0.1 mg/mL streptomycin, and 1% non-essential amino acids (Merck) in a humidified incubator at 5% CO2 and 37 °C. Calu-3suggested: KCLB Cat# 30055, RRID:CVCL_0609)After 48 hours of treatment, cell viability was evaluated by adding MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (Sigma) to the cells for 4 h and OD562 measurements according to the manufacturer’s protocols (Sigma). 2.3 Inoculation of cells and drug treatment: For infection, polarized Calu-3 and Vero cells were washed with PBS and inoculated at a multiplicity of infection (MOI) of 0.1 (Calu-3) or 0.01 (Vero E6) of virus diluted in infection-PBS (containing 0.2% BSA, 1% CaCl2, 1% MgCl2, 100 U/mL penicillin and 0.1 mg/mL streptomycin) at 37°C for 1 hours. Verosuggested: NoneVero E6suggested: NoneSoftware and Algorithms Sentences Resources Virus plaques were visualized by staining with neutral red, and virus titers were calculated as plaque-forming units (PFU) per mL. 2.5 Data and statistical analysis: A priori power analysis using G*Power 3.1 (Faul et al., 2007)) was performed to determine the sample sizes required to detect > 90% reduction in virus titers at powers > 0.8. G*Powersuggested: (G*Power, RRID:SCR_013726)Data were analyzed using the software GraphPad Prism version 8.00 (GraphPad). GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)Drug combinatory effects were analyzed by using SynergyFinder, an open-source free stand-alone web application for the analysis of drug combination data (Ianevski et al., 2017). SynergyFindersuggested: (SynergyFinder, RRID:SCR_019318)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 found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
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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