My Journey to a CURE
  • Welcome
  • Special Thanks
  • Blog
  • My Favorite Products
  • Contact Email
  • Free Motivational & Inspirational E-Books
  • Magic of Magnesium
  • Johns Journey to a Prostate Cure
  • CS BrainSense
  • Wifi Router Guard
  • Smart Meter Guard
  • VIBE Resance Frequency Therapy

Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro

3/10/2020

 
Picture
Dear Editor,
In December 2019, a novel pneumonia caused by a previously unknown pathogen emerged in Wuhan, a city of 11 million people in central China. The initial cases were linked to exposures in a seafood market in Wuhan.1 As of January 27, 2020, the Chinese authorities reported 2835 confirmed cases in mainland China, including 81 deaths. Additionally, 19 confirmed cases were identified in Hong Kong, Macao and Taiwan, and 39 imported cases were identified in Thailand, Japan, South Korea, United States, Vietnam, Singapore, Nepal, France, Australia and Canada. The pathogen was soon identified as a novel coronavirus (2019-nCoV), which is closely related to sever acute respiratory syndrome CoV (SARS-CoV).2 

​Currently, there is no specific treatment against the new virus. Therefore, identifying effective antiviral agents to combat the disease is urgently needed

An efficient approach to drug discovery is to test whether the existing antiviral drugs are effective in treating related viral infections. The 2019-nCoV belongs to Betacoronavirus which also contains SARS-CoV and Middle East respiratory syndrome CoV (MERS-CoV). Several drugs, such as ribavirin, interferon, lopinavir-ritonavir, corticosteroids, have been used in patients with SARS or MERS, although the efficacy of some drugs remains controversial.3 

​In this study, we evaluated the antiviral efficiency of five FAD-approved drugs including ribavirin, penciclovir, nitazoxanide, nafamostat, chloroquine and two well-known broad-spectrum antiviral drugs remdesivir (GS-5734) and favipiravir (T-705) against a clinical isolate of 2019-nCoV in vitro.

Standard assays were carried out to measure the effects of these compounds on the cytotoxicity, virus yield and infection rates of 2019-nCoVs. Firstly, the cytotoxicity of the candidate compounds in Vero E6 cells (ATCC-1586) was determined by the CCK8 assay. Then, Vero E6 cells were infected with nCoV-2019BetaCoV/Wuhan/WIV04/20192 at a multiplicity of infection (MOI) of 0.05 in the presence of varying concentrations of the test drugs. DMSO was used in the controls. Efficacies were evaluated by quantification of viral copy numbers in the cell supernatant via quantitative real-time RT-PCR (qRT-PCR) and confirmed with visualization of virus nucleoprotein (NP) expression through immunofluorescence microscopy at 48 h post infection (p.i.) (cytopathic effect was not obvious at this time point of infection). Among the seven tested drugs, high concentrations of three nucleoside analogs including ribavirin (half-maximal effective concentration (EC50) = 109.50 μM, half-cytotoxic concentration (CC50) > 400 μM, selectivity index (SI) > 3.65), penciclovir (EC50 = 95.96 μM, CC50 > 400 μM, SI > 4.17) and favipiravir (EC50 = 61.88 μM, CC50 > 400 μM, SI > 6.46) were required to reduce the viral infection (Fig. 1a and Supplementary information, Fig. S1). However, favipiravir has been shown to be 100% effective in protecting mice against Ebola virus challenge, although its EC50 value in Vero E6 cells was as high as 67 μM,4 suggesting further in vivo studies are recommended to evaluate this antiviral nucleoside. Nafamostat, a potent inhibitor of MERS-CoV, which prevents membrane fusion, was inhibitive against the 2019-nCoV infection (EC50 = 22.50 μM, CC50 > 100 μM, SI > 4.44). Nitazoxanide, a commercial antiprotozoal agent with an antiviral potential against a broad range of viruses including human and animal coronaviruses, inhibited the 2019-nCoV at a low-micromolar concentration (EC50 = 2.12 μM; CC50 > 35.53 μM; SI > 16.76). Further in vivo evaluation of this drug against 2019-nCoV infection is recommended. Notably, two compounds remdesivir (EC50 = 0.77 μM; CC50 > 100 μM; SI > 129.87) and chloroquine (EC50 = 1.13 μM; CC50 > 100 μM, SI > 88.50) potently blocked virus infection at low-micromolar concentration and showed high SI (Fig. 1a, b).

​Fig. 1: The antiviral activities of the test drugs against 2019-nCoV in vitro.

a Vero E6 cells were infected with 2019-nCoV at an MOI of 0.05 in the treatment of different doses of the indicated antivirals for 48 h. The viral yield in the cell supernatant was then quantified by qRT-PCR. Cytotoxicity of these drugs to Vero E6 cells was measured by CCK-8 assays. The left and right Y-axis of the graphs represent mean % inhibition of virus yield and cytotoxicity of the drugs, respectively. The experiments were done in triplicates. b Immunofluorescence microscopy of virus infection upon treatment of remdesivir and chloroquine. Virus infection and drug treatment were performed as mentioned above. At 48 h p.i., the infected cells were fixed, and then probed with rabbit sera against the NP of a bat SARS-related CoV2 as the primary antibody and Alexa 488-labeled goat anti-rabbit IgG (1:500; Abcam) as the secondary antibody, respectively. The nuclei were stained with Hoechst dye. Bars, 100 μm. c and d Time-of-addition experiment of remdesivir and chloroquine. For “Full-time” treatment, Vero E6 cells were pre-treated with the drugs for 1 h, and virus was then added to allow attachment for 2 h. Afterwards, the virus–drug mixture was removed, and the cells were cultured with drug-containing medium until the end of the experiment. For “Entry” treatment, the drugs were added to the cells for 1 h before viral attachment, and at 2 h p.i., the virus–drug mixture was replaced with fresh culture medium and maintained till the end of the experiment. For “Post-entry” experiment, drugs were added at 2 h p.i., and maintained until the end of the experiment. For all the experimental groups, cells were infected with 2019-nCoV at an MOI of 0.05, and virus yield in the infected cell supernatants was quantified by qRT-PCR c and NP expression in infected cells was analyzed by Western blot d at 14 h p.i.

Remdesivir has been recently recognized as a promising antiviral drug against a wide array of RNA viruses (including SARS/MERS-CoV5) infection in cultured cells, mice and nonhuman primate (NHP) models. It is currently under clinical development for the treatment of Ebola virus infection.6 Remdesivir is an adenosine analogue, which incorporates into nascent viral RNA chains and results in pre-mature termination.7 Our time-of-addition assay showed remdesivir functioned at a stage post virus entry (Fig. 1c, d), which is in agreement with its putative anti-viral mechanism as a nucleotide analogue. Warren et al. showed that in NHP model, intravenous administration of 10 mg/kg dose of remdesivir resulted in concomitant persistent levels of its active form in the blood (10 μM) and conferred 100% protection against Ebola virus infection.7 Our data showed that EC90 value of remdesivir against 2019-nCoV in Vero E6 cells was 1.76 μM, suggesting its working concentration is likely to be achieved in NHP. Our preliminary data (Supplementary information, Fig. S2) showed that remdesivir also inhibited virus infection efficiently in a human cell line (human liver cancer Huh-7 cells), which is sensitive to 2019-nCoV.2

Chloroquine, a widely-used anti-malarial and autoimmune disease drug, has recently been reported as a potential broad-spectrum antiviral drug.8,9 Chloroquine is known to block virus infection by increasing endosomal pH required for virus/cell fusion, as well as interfering with the glycosylation of cellular receptors of SARS-CoV.10 Our time-of-addition assay demonstrated that chloroquine functioned at both entry, and at post-entry stages of the 2019-nCoV infection in Vero E6 cells (Fig. 1c, d). Besides its antiviral activity, chloroquine has an immune-modulating activity, which may synergistically enhance its antiviral effect in vivo. Chloroquine is widely distributed in the whole body, including lung, after oral administration. The EC90 value of chloroquine against the 2019-nCoV in Vero E6 cells was 6.90 μM, which can be clinically achievable as demonstrated in the plasma of rheumatoid arthritis patients who received 500 mg administration.11 Chloroquine is a cheap and a safe drug that has been used for more than 70 years and, therefore, it is potentially clinically applicable against the 2019-nCoV.

Our findings reveal that remdesivir and chloroquine are highly effective in the control of 2019-nCoV infection in vitro. Since these compounds have been used in human patients with a safety track record and shown to be effective against various ailments, we suggest that they should be assessed in human patients suffering from the novel coronavirus disease.


References
  1. 1.Huang, C. L. et al. The Lancet https://doi.org/10.1016/S0140-6736(20)30183-5 (2020).
    • CAS
    • Article
    • Google Scholar
  2. 2.Zhou, P. et al. Nature (accepted).
  3. 3.Zumla, A., Chan, J. F., Azhar, E. I., Hui, D. S. & Yuen, K. Y. Nat. Rev. Drug Discov. 15, 327–347 (2016).
    • CAS
    • Article
    • Google Scholar
  4. 4.Oestereich, L. et al. Antivir. Res. 105, 17–21 (2014).
    • CAS
    • Article
    • Google Scholar
  5. 5.Sheahan, T. P. et al. Sci. Transl. Med. 9, eaal3653 (2017).
    • Article
    • Google Scholar
  6. 6.Mulangu, S. et al. N. Engl. J. Med. 381, 2293–2303 (2019).
    • CAS
    • Article
    • Google Scholar
  7. 7.Warren, T. K. et al. Nature 531, 381–385 (2016).
    • CAS
    • Article
    • Google Scholar
  8. 8.Savarino, A., Di Trani, L., Donatelli, I., Cauda, R. & Cassone, A. Lancet Infect. Dis. 6, 67–69 (2006).
    • Article
    • Google Scholar
  9. 9.Yan, Y. et al. Cell Res. 23, 300–302 (2013).
    • CAS
    • Article
    • Google Scholar
  10. 10.Vincent, M. J. et al. Virol. J. 2, 69 (2005).
    • Article
    • Google Scholar
  11. 11.Mackenzie, A. H. Am. J. Med. 75, 40–45 (1983).
    • CAS
    • Article
    • Google Scholar
Download references
AcknowledgementsWe thank Xi Wang, Yan Wu, Weijuan Shang, Huanyu Zhang, Yufeng Li, Hengrui Hu, Xiaming Jiang, Yuan Sun, from Wuhan Institute of Virology for their essential assistance with this study. We thank Prof. Fei Deng from National Virus Resource Center, and Tao Du, Jia Wu and Hao Tang from BSL-3 Laboratory of Wuhan Institute of Virology for their critical support. We thank Prof. Yanyi Wang and other colleagues of Wuhan Institute of Virology and Wuhan National Biosafety Laboratory for their excellent coordination. We thank Dr. Basil Arif for scientific editing of the manuscript. We thank the anonymous reviewers for their valuable suggestions. This work was supported in part by grants from the National Science and Technology Major Projects for “Major New Drugs Innovation and Development” (directed by Prof. Song Li) (2018ZX09711003), the National Natural Science Foundation of China (31621061), and the Emergency Scientific Research Project for 2019-nCoV from Hubei Province (to Profs. Zhengli Shi and Gengfu Xiao).

Author information
Author notes
  1. These authors contributed equally: Manli Wang, Ruiyuan Cao, Leike Zhang, Xinglou Yang.
Affiliations
  1. State Key Laboratory of Virology, Wuhan Institute of Virology, Center for Biosafety Mega-Science, Chinese Academy of Sciences, 430071, Wuhan, China
    • Manli Wang
    • , Leike Zhang
    • , Xinglou Yang
    • , Jia Liu
    • , Mingyue Xu
    • , Zhengli Shi
    • , Zhihong Hu
    •  & Gengfu Xiao
  2. National Engineering Research Center for the Emergency Drug, Beijing Institute of Pharmacology and Toxicology, 100850, Beijing, China
    • Ruiyuan Cao
    •  & Wu Zhong
Authors
  1. Manli Wang
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  2. Ruiyuan Cao
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  3. Leike Zhang
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  4. Xinglou Yang
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  5. Jia Liu
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  6. Mingyue Xu
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  7. Zhengli Shi
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  8. Zhihong Hu
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  9. Wu Zhong
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar
  10. Gengfu Xiao
    View author publications
    You can also search for this author in
    • PubMed
    • Google Scholar

.X., W.Z., Z.H., M.W., R.C., and L.Z. conceived and designed the experiments. X.Y., J.L., M.X., M.W., R.C., and L.Z. participated in multiple experiments; G.X., W.Z., Z.H., Z.S., M.W., R.C., and L.Z. analyzed the data. M.W., L.Z., R.C., and Z.H. wrote the manuscript. G.X., W.Z., and Z.H. provided the final approval of the manuscript.

Corresponding authorsCorrespondence to Zhihong Hu or Wu Zhong or Gengfu Xiao.
Ethics declarationsCompeting interestsThe authors declare no competing interests.
Supplementary informationSupplementary information, Materials and FiguresRights and permissionsOpen Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro

Author: Manli Wang et al

Publication: Cell Research

Publisher:Springer Nature

Date:Feb 4, 2020
Copyright © 2020, Springer Nature
Creative Commons
This is an open access article distributed under the terms of the Creative Commons CC BY license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

You are not required to obtain permission to reuse this article.
To request permission for a type of use not listed, please contact Springer Nature

https://www.nature.com/articles/s41422-020-0282-0


This is only available by prescription from your doctor.

Comments are closed.

    Categories

    All
    Adaptogen
    Air Purification
    Alkalinize PH
    Allergy
    Aloe Vera
    ALS
    Alzheimers
    Amazon
    Amino Acids
    Amish Origins Pain Relief Cream
    Anti Aging
    Antioxidant
    ANXIETY
    Apple Cider Vinegar
    Arginine
    Arthritis
    Artificial-sweeteners
    Aspartame
    Asthma
    Auto Immune
    B-12
    Back Pain
    Bacteria
    Baking Soda
    Balance
    Beet Root Super Powder
    Berberine
    Beta Glucan
    BioDefense
    BioReign
    Bioreigns Cbd
    Birth Defects
    Black-cumin
    Black-seed-oil
    Bloating
    Blood-flow
    Blood-flow
    Blood Glucose
    Blood Sugar
    Bone Loss
    Books
    Bowel Function
    Brain Function
    Brain Mental Vitality
    Brain Sense
    Breast Cancer
    Broccoli
    Burns
    Calcium
    Cancer
    Cancer-awareness-jewelry
    Cancer-treatments
    Cardiovascular
    Carrot Powder Organic
    Cayenne Pepper
    CBD Hemp Organic
    CBD - Hemp - Organic
    Cell Fuzion
    Cell Phone
    Cell Phone Radiation
    Cellular Hydrate
    Chelation
    Chloride
    Chlorine Shower
    Chloroquine
    Cholestrol
    Cleaning
    Cleaning Fruits & Produce
    Cleanses
    Cleansing
    Collagen
    Colon
    Colonel Joe
    Computer Back Up
    Congestion
    Corey's Digs
    Coronavirus
    Corpus Christi Water Contamination
    Cortisol
    Covid 19 Virus
    Cramps
    CS BrainSense
    Curcumin
    Cyanobacteria
    D3 And K2
    Dandelion Root
    DEFEND Super Patch
    Defense
    Dehydration
    Dementia
    Depression
    Detox
    DIABETES
    Diet
    Digestion
    Digestive
    Disease Prevention
    Disinfectant
    Dna Repair
    Dog Monitor Camera
    DOGS
    Drug Free
    Earthing
    Elderberry
    Electrolytes
    EMF Protection
    EMR Defender
    Energy
    Environment Friendly
    Enzymes
    Essential Oils
    Extra Virgin Olive Oil
    Fatigue
    Fatty Liver
    FDA
    Fish Oil
    Florida Eco Disaster
    Flow Super Patch
    Flu
    Fluoride
    Focus
    Focus Super Patch
    Freedom Super Patch
    Free E Books
    Free Mart Products
    Frequencies
    Fulvic Humic Acid
    Germs
    Global Healing
    Glycine
    Glyphosate
    Grape-seed-extract
    Greens
    Green Tea
    Green-tea
    Grounding
    Groupon
    Gut Health
    Hair And Nails
    Hair Loss Growth
    Hand Sanitizer DIY\
    Hand Wipes - DIY
    Hart Procedure
    Headaches
    Healthy Hair And Nails
    Healthy Joints
    Healthy Skin
    Heart Disease
    Heart Health
    Herbicides
    High Blood Pressure
    Holidays
    Homeostatis
    Huperzine-hupa
    Hydrate
    Hydration
    Hydroxychloroquine
    Hypertension
    Immune Support
    Immune System
    Immune-system
    Immunotherapy
    Inflammation
    Inspiration
    Iodine
    Johns Journey
    Joint Pain
    Kidney Disease
    Lemons
    Liberty Super Patch
    Limitless
    Liver
    Liver Cleanse
    Longevity
    Lou Genrigs Disease
    L Serine
    L-serine
    Lung
    Lycopene
    Magnesium
    Meal Replacement
    Memory
    Minerals
    MiracleII
    Mobility
    Monsanto
    Moringa
    MS
    Muscadine
    Muscadine Grape
    Muscadine Seed Oil
    Muscle Tissue
    Mushrooms
    My Story
    Nano Soma
    Natural Fertilizer
    Natural Glow Hydration
    Nervous System
    Neuropathy
    Nigella Sativa
    Now We No Movement
    Nutrition
    Off The Grid
    Olive Leaf Extract
    Omega 3
    On The Go Knowledge Pantry
    Oregano
    Organic Health
    Organic Herbal Extracts
    Organic Teas
    ORGANOPHOSPHATE
    Osteoarthritis
    Oxygen
    Pain Relief
    Pancreatic Cancer
    Parkinsons
    PEMF
    Pesticides
    Pet Health
    Pets
    Poisoning
    Pollen
    Pomegranate
    Potassium
    Prebiotics
    Precious Prills
    Premature Aging
    Preparation
    Probiotic
    Prostate Cancer
    Protein
    Quercetin
    Radiation
    Rashes
    Raspex
    Red Tide Illness
    Red Tide Illness Remedies
    Red Tide TOXIC BACTERIA
    Remdesivir
    Resona
    Respiratory
    Restore
    Resveratrol
    Reverse Osmosis
    RFT
    Sage
    Sale
    Sanitary Napkins
    Saving Money (Earning $)
    Selenium
    Sept 11
    Shark Cartilage
    Shiaqga
    Shilajit
    Shopping
    Shower Filter
    Siahus
    Siesta Key Red Tide TOXIC Algae
    Silver
    Sinus
    SKIN CARE
    Skin Sorcery
    Sleep
    Sleep REM Super Patch
    Slver
    Soap Ans Laundry
    Special
    Special Thanks
    Sponsors Movies Websites
    Sports Performance
    Stem Cell
    Strength
    Stress
    Stroke
    Sunburn
    Sunscreen
    Superfoods
    Super Patch Company
    Supplement Discounts
    TAKE ACTION
    Taurine
    Testimonial
    Throat Pain
    Thyroid
    Thyroid Health
    Toothpaste
    Total Wellness
    Toxic Metals
    Toxin Removal
    Toxins
    True Hydrate
    Turmeric
    Ultrapure Water
    Veterans
    VIBE
    Victory SUPER Patch
    Vitamin B12
    Vitamin C
    Vitamin D3
    Vitamin K2
    Washing Your Hands
    Water
    Water Devices Treatments
    Water Filtration
    Weed Control
    Weight Loss
    Welcome
    Wrinkle Removal
    Wrinkles
    Zeolite
    Zeolite DHQ
    Zeotrex
    ZeroWater
    Zinc
    ZNatural

    Archives

    March 2025
    February 2025
    January 2025
    August 2024
    June 2024
    May 2024
    April 2024
    March 2024
    January 2024
    November 2023
    October 2023
    September 2023
    July 2023
    June 2023
    May 2023
    April 2023
    January 2023
    October 2022
    September 2022
    August 2022
    June 2022
    May 2022
    April 2022
    December 2021
    November 2021
    September 2021
    August 2021
    May 2021
    April 2021
    January 2021
    December 2020
    July 2020
    May 2020
    April 2020
    March 2020
    February 2020
    January 2020
    December 2019
    November 2019
    October 2019
    September 2019
    August 2019
    July 2019
    May 2019
    March 2019
    January 2019
    December 2018
    November 2018
    October 2018
    September 2018
    August 2018
    July 2018
    June 2018
    April 2018
    March 2018
    February 2018
    January 2018
    December 2017
    November 2017
    October 2017
    August 2017
    July 2017
    May 2017
    April 2017
    February 2017
    January 2017
    December 2016
    November 2016
    October 2016
    September 2016
    August 2016
    July 2016
    June 2016
    May 2016
    January 2016
    December 2015
    November 2015
    October 2015
    September 2015
    August 2015
    July 2015
    February 2015
    July 2014
    June 2014
    November 2011

    RSS Feed

    Picture
    Picture
    Picture
    Picture
    Zeotrex® is an herbal blend that helps rejuvenate vitality, energy, mental clarity, and overall wellness through the detoxification of chemicals and metals.
    Pura Vida Best Sellers
    Picture
    Picture
    Picture
    Picture
    Picture
    Click to set custom HTML


    Fluoride Filter
    11 Free Tips to Reduce Your Fluoride Exposure
    Picture
    Save up to 33% with Bundle Deals -  Shop Now
    Phenoh
My Journey to A Cure Copyright 2010 - 2024