Repurposing the Ebola and Marburg Virus Inhibitors Tilorone, Quinacrine, and Pyronaridine: In Vitro Activity against SARS-CoV‑2 and Potential Mechanisms
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Abstract
Severe acute respiratory coronavirus 2 (SARS-CoV-2) is a newly identified virus that has resulted in over 2.5 million deaths globally and over 116 million cases globally in March, 2021. Small-molecule inhibitors that reverse disease severity have proven difficult to discover. One of the key approaches that has been widely applied in an effort to speed up the translation of drugs is drug repurposing. A few drugs have shown in vitro activity against Ebola viruses and demonstrated activity against SARS-CoV-2 in vivo. Most notably, the RNA polymerase targeting remdesivir demonstrated activity in vitro and efficacy in the early stage of the disease in humans. Testing other small-molecule drugs that are active against Ebola viruses (EBOVs) would appear a reasonable strategy to evaluate their potential for SARS-CoV-2. We have previously repurposed pyronaridine, tilorone, and quinacrine (from malaria, influenza, and antiprotozoal uses, respectively) as inhibitors of Ebola and Marburg viruses in vitro in HeLa cells and mouse-adapted EBOV in mice in vivo. We have now tested these three drugs in various cell lines (VeroE6, Vero76, Caco-2, Calu-3, A549-ACE2, HUH-7, and monocytes) infected with SARS-CoV-2 as well as other viruses (including MHV and HCoV 229E). The compilation of these results indicated considerable variability in antiviral activity observed across cell lines. We found that tilorone and pyronaridine inhibited the virus replication in A549-ACE2 cells with IC50 values of 180 nM and IC50 198 nM, respectively. We used microscale thermophoresis to test the binding of these molecules to the spike protein, and tilorone and pyronaridine bind to the spike receptor binding domain protein with Kd values of 339 and 647 nM, respectively. Human Cmax for pyronaridine and quinacrine is greater than the IC50 observed in A549-ACE2 cells. We also provide novel insights into the mechanism of these compounds which is likely lysosomotropic.
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SciScore for 10.1101/2020.12.01.407361: (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. Table 2: Resources
No key resources detected.
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: We detected the following sentences addressing limitations in the study:Despite this limitation, valuable information about the virus infection and replication can be learned from studies using these cell lines. A previous study (Chu et al., 2020) assessed 25 cell lines derived from different tissues …
SciScore for 10.1101/2020.12.01.407361: (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. Table 2: Resources
No key resources detected.
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: We detected the following sentences addressing limitations in the study:Despite this limitation, valuable information about the virus infection and replication can be learned from studies using these cell lines. A previous study (Chu et al., 2020) assessed 25 cell lines derived from different tissues or organs and host species and reported that cytopathic effects were only seen in VeroE6 and FRhK4 cells after SARS-CoV-2 inoculation for up to 120 hpi. These findings are important for optimization of antiviral assays based on cell protection assessment, because cell lines without obvious cytopathic effects might lead to overestimation of cell viability and drug efficacy (Chu et al., 2020). For efficient SARS-CoV-2 research, a cell line, such as Vero cells, that can easily replicate and isolate the virus is essential, but they have been shown not to produce interferon (IFN) when infected with Newcastle disease virus, rubella virus, and other viruses (Desmyter et al., 1968). Under previously described experimental conditions, productive SARS-CoV-2 replication in A549 cells was erratic (Sacramento et al., 2020) which can be overcome by preparing A549 cells overexpressing ACE2 (as used in the current study). The differences in responses in different cell lines could be accounted for by the basic biochemistry, for example hepatic cells, like Huh-7, are equipped with enzymes to synthesize nucleotides, carbohydrates and lipids (Nwosu et al., 2018). Hence it is not surprising that the highest potency of remdesivir against SARS-CoV-2 is found in Huh-7 cells...
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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