Identification of Anti-Severe Acute Respiratory Syndrome-Related Coronavirus 2 (SARS-CoV-2) Oxysterol Derivatives In Vitro
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (ScreenIT)
Abstract
The development of effective antiviral drugs targeting the severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) is urgently needed to combat the coronavirus disease 2019 (COVID-19). We have previously studied the use of semi-synthetic derivatives of oxysterols, oxidized derivatives of cholesterol as drug candidates for the inhibition of cancer, fibrosis, and bone regeneration. In this study, we screened a panel of naturally occurring and semi-synthetic oxysterols for anti-SARS-CoV-2 activity using a cell culture infection assay. We show that the natural oxysterols, 7-ketocholesterol, 22(R)-hydroxycholesterol, 24(S)-hydroxycholesterol, and 27-hydroxycholesterol, substantially inhibited SARS-CoV-2 propagation in cultured cells. Among semi-synthetic oxysterols, Oxy210 and Oxy232 displayed more robust anti-SARS-CoV-2 activities, reducing viral replication more than 90% at 10 μM and 99% at 15 μM, respectively. When orally administered in mice, peak plasma concentrations of Oxy210 fell into a therapeutically relevant range (19 μM), based on the dose-dependent curve for antiviral activity in our cell-based assay. Mechanistic studies suggest that Oxy210 reduced replication of SARS-CoV-2 by disrupting the formation of double-membrane vesicles (DMVs); intracellular membrane compartments associated with viral replication. Our study warrants further evaluation of Oxy210 and Oxy232 as a safe and reliable oral medication, which could help protect vulnerable populations with increased risk of developing COVID-19.
Article activity feed
-
-
SciScore for 10.1101/2021.01.31.429001: (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 Detection of viral N protein: Viral N protein was detected using a rabbit anti-SARS-CoV N antibody [32] as a primary antibody with AlexaFluor 568 anti-rabbit IgG or anti-rabbit IgG-HRP (Thermo Fisher) as secondary antibodies together with DAPI to stain the nucleus by indirect immunofluorescence as described previously [33]. anti-SARS-CoV Nsuggested: Noneanti-rabbit IgGsuggested: Noneanti-rabbit IgG-HRPsuggested: NoneExperimental Models: Cell Lines Sentences R… SciScore for 10.1101/2021.01.31.429001: (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 Detection of viral N protein: Viral N protein was detected using a rabbit anti-SARS-CoV N antibody [32] as a primary antibody with AlexaFluor 568 anti-rabbit IgG or anti-rabbit IgG-HRP (Thermo Fisher) as secondary antibodies together with DAPI to stain the nucleus by indirect immunofluorescence as described previously [33]. anti-SARS-CoV Nsuggested: Noneanti-rabbit IgGsuggested: Noneanti-rabbit IgG-HRPsuggested: NoneExperimental Models: Cell Lines Sentences Resources Cell culture: VeroE6/TMPRSS2 cells, VeroE6 cells overexpressing transmembrane protease, serine 2 (TMPRSS2) [20, 29], were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Wako) supplemented with 10% fetal bovine serum (FBS; Cell Culture Bioscience), 10 units/mL penicillin, 10 μg/mL streptomycin, 10 mM HEPES (pH 7.4), and 1 mg/mL G418 (Nacalai) at 37°C in 5% CO2. VeroE6suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)HepG2-hNTCP-C4 cells, a HepG2 cell clone overexpressing the HDV entry receptor, sodium taurocholate cotrasporting polypeptide (NTCP), and highly susceptible to HDV infection [6] were cultured in GlutaMax (Invitrogen) supplemented with 10 units/ml penicillin, 10 μg/ml streptomycin, 10% FBS, 10 mM HEPES (pH 7.4), 50 μM hydrocortisone, and 5 μg/ml insulin at 37°C in 5% CO2. HepG2suggested: NoneVeroE6/TMPRSS2 cells were inoculated with SARS-CoV-2 at an MOI of 0.001 (Figure 1B, 1C, 2A, and 2B), 0.003 (Figure 1D, 2C, and 3A), and 1 (Figure 3B) for 1 h and unbound virus removed by washing. VeroE6/TMPRSS2suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)For examination of Hh activity, NIH3T3 cells pretreated with the compounds for 2 h were treated with conditioned medium from CAPAN-1 human pancreatic tumor cells that contain Shh in the absence or presence of the compounds. NIH3T3suggested: KCB Cat# KCB 200645YJ, RRID:CVCL_0594)Hepatitis D virus (HDV) replication assay: HDV were recovered from the culture supernatant of Huh7 cells transfected with the plasmids for HDV genome and for hepatitis B virus surface antigen [34]. Huh7suggested: CLS Cat# 300156/p7178_HuH7, RRID:CVCL_0336)HepG2-hNTCP-C4 cells were inoculated with HDV for 16 h and were further cultured for 6 days in the presence or absence of Oxy210 to detect intracelular HDV RNA [34]. HepG2-hNTCP-C4suggested: NoneResults 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 scite Reference Check: We found no unreliable references.
-
