Prophylactic and Therapeutic Inhibition of In Vitro SARS-CoV-2 Replication by Oleandrin
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Abstract
With continued expansion of the COVID-19 pandemic, antiviral drugs are desperately needed to treat patients at high risk of life-threatening disease and even to limit spread if administered early during infection. Typically, the fastest route to identifying and licensing a safe and effective antiviral drug is to test those already shown safe in early clinical trials for other infections or diseases. Here, we tested in vitro oleandrin, derived from the Nerium oleander plant and shown previously to have inhibitory activity against several viruses. Using Vero cells, we found that prophylactic oleandrin administration at concentrations down to 0.05 μg/ml exhibited potent antiviral activity against SARS-CoV-2, with an 800-fold reduction in virus production, and a 0.1 μg/ml dose resulted in a greater than 3,000-fold reduction in infectious virus production. The EC 50 values were 11.98ng/ml when virus output was measured at 24 hours post-infection, and 7.07ng/ml measured at 48 hours post-infection. Therapeutic (post-infection) treatment up to 24 hours after infection of Vero cells also reduced viral titers, with the 0.1 μg/ml dose causing greater than 100-fold reductions as measured at 48 hours, and the 0.05 μg/ml dose resulting in a 78-fold reduction. The potent prophylactic and therapeutic antiviral activities demonstrated here strongly support the further development of oleandrin to reduce the severity of COVID-19 and potentially also to reduce spread by persons diagnosed early after infection.
IMPORTANCE
COVID-19, a pandemic disease caused by infection with SARS-CoV-2, has swept around the world to cause millions of infections and hundreds-of-thousands of deaths due to the lack of vaccines and effective therapeutics. We tested oleandrin, derived from the Nerium oleander plant and shown previously to reduce the replication of several viruses, against SARS-CoV-2 infection of Vero cells. When administered both before and after virus infection, nanogram doses of oleandrin significantly inhibited replication by up to 3,000-fold, indicating the potential to prevent disease and virus spread in persons recently exposed to SARS-CoV-2, as well as to prevent severe disease in persons at high risk. These results indicate that oleandrin should be tested in animal models and in humans exposed to infection to determine its medical usefulness in controlling the pandemic.
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SciScore for 10.1101/2020.07.15.203489: (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
Experimental Models: Cell Lines Sentences Resources Vero CCL81 cells were seeded, infected, washed, and returned to the incubator as before. Vero CCL81suggested: NonePlaque Assay: Vero E6 cells were seeded in 6-well plates (Thermo Scientific, Waltham, MA) to a confluency of approximately 90%. Vero E6suggested: NoneSoftware and Algorithms Sentences Resources Absorbance was measured at 490nm and 680nm on a Synergy HT plate reader (BioTek, Winooski, VT) with Gen5 data analysis software, … SciScore for 10.1101/2020.07.15.203489: (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
Experimental Models: Cell Lines Sentences Resources Vero CCL81 cells were seeded, infected, washed, and returned to the incubator as before. Vero CCL81suggested: NonePlaque Assay: Vero E6 cells were seeded in 6-well plates (Thermo Scientific, Waltham, MA) to a confluency of approximately 90%. Vero E6suggested: NoneSoftware and Algorithms Sentences Resources Absorbance was measured at 490nm and 680nm on a Synergy HT plate reader (BioTek, Winooski, VT) with Gen5 data analysis software, version 2.05 (BioTek, Winooski, VT). Gen5suggested: (Gen5, RRID:SCR_017317)All graphs were generated in Prism version 7.03 (GraphPad, San Diego, CA). Prismsuggested: (PRISM, RRID:SCR_005375)GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)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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SciScore for 10.1101/2020.07.15.203489: (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
Experimental Models: Cell Lines Sentences Resources Therapeutic (post-infection) treatment up to 24 hours after infection of Vero cells also reduced viral titers, with the 0.1 µg/ml dose causing greater than 100-fold reductions as measured at 48 hours, and the 0.05 µg/ml dose resulting in a 78-fold reduction. Verosuggested: NoneMethods Cells, Viruses, and Oleandrin Vero CCL81 cells were used for the prophylactic … SciScore for 10.1101/2020.07.15.203489: (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
Experimental Models: Cell Lines Sentences Resources Therapeutic (post-infection) treatment up to 24 hours after infection of Vero cells also reduced viral titers, with the 0.1 µg/ml dose causing greater than 100-fold reductions as measured at 48 hours, and the 0.05 µg/ml dose resulting in a 78-fold reduction. Verosuggested: NoneMethods Cells, Viruses, and Oleandrin Vero CCL81 cells were used for the prophylactic and therapeutic assays (ATCC, Manassas, VA) Vero CCL81suggested: NonePlaque assays were performed in Vero E6 cells, kindly provided by Vineet Menachery (UTMB, Galveston, TX). Vero E6suggested: CVCL_XD71Software and Algorithms Sentences Resources Thus, considering the beneficial anti-inflammatory activity of Nerium oleander extracts combined with the suggestion that oleandrin might protect against neurological deficits, Phoenix Biotechnology, Inc. (San Antonio, TX) has developed an extract for sublingual administration that it will quickly explore for efficacy in appropriate in vivo studies. Phoenix Biotechnologysuggested: NoneAbsorbance was measured at 490nm and 680nm on a Synergy HT plate reader (BioTek, Winooski, VT) with Gen5 data analysis software, version 2.05 (BioTek, Winooski, VT). Gen5suggested: (Gen5, SCR_017317)All graphs were generated in Prism version 7.03 (GraphPad, San Diego, CA). Prismsuggested: (PRISM, SCR_005375)<div style="margin-bottom:8px"> <div><b>GraphPad</b></div> <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div> </div> </td></tr></table>Data from additional tools added to each annotation on a weekly basis.
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