Lipid nanoparticle formulation of niclosamide (nano NCM) effectively inhibits SARS-CoV-2 replication in vitro

  1. Guankui Wang
  2. Hanmant Gaikwad
  3. Mary McCarthy
  4. Mercedes Gonzalez-Juarrero
  5. Yue Li
  6. Michael Armstrong
  7. Nichole Reisdorph
  8. Thomas Morrison
  9. Dmitri Simberg

Reviewed by

Read the full article

Listed in

Log in to save this article

Abstract

<img src=” https://s3.amazonaws.com/production.scholastica/article/18813/large/prnano_612020.jpg?1610907930”>

As exemplified by the COVID-19 pandemic, highly infectious respiratory viruses can spread rapidly in the population because of a lack of effective approaches to control viral replication and spread. Niclosamide is an old anthelminthic drug (see the essential medicine list of the World Health Organization) with pleiotropic pharmacological activities. Several recent publica-tions demonstrated that niclosamide has broad antiviral activities and potently inhibits viral rep-lication, including replication of SARS-CoV-2, SARS-CoV, and dengue viruses. Unfortunately, it is almost entirely insoluble in water, which limits its clinical use. We developed a cost-effective lipid nanoparticle formulation of niclosamide (nano-NCM) using only Food and Drug Admin-istration-approved excipient and demonstrated potency against SARS-CoV-2 infection in Vero E6 and ACE2-expressing lung epithelium cells in vitro.

Article activity feed

  1. Version published to 10.33218/001c.18813
  2. ScreenIT

    SciScore for 10.1101/2020.12.18.423509: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Cells were fixed with 4% paraformaldehyde and probed with 1,000 ng/mL of an anti-SARS-CoV-2 spike monoclonal antibody (CR3022, Absolute Antibody) in Perm Wash (1X PBS/0.1% saponin).
    anti-SARS-CoV-2
    suggested: (Abcam Cat# ab273074, RRID:AB_2847846)
    CR3022
    suggested: (Imported from the IEDB Cat# CR3022, RRID:AB_2848080)
    Experimental Models: Cell Lines
    SentencesResources
    Human ACE2 (angiotensin-converting enzyme 2) stably transfected A549 cells (hACE2-A549) were obtained from Dr. Mario Santiago Laboratory at CU Anschutz.
    A549
    suggested: None
    Cells were grown in 5% CO2 atmosphere at 37 °C in DMEM containing 10% FBS, 100 U/mL penicillin and 100 ng/mL streptomycin for Vero E6 cells or puromycin (0.5 µg/mL) for hACE2A549 cells.
    hACE2A549
    suggested: None
    Cytotoxicity (CC50) to the noninfected Vero E6 and hACE2-A549 cells was measured with MTT assay.
    hACE2-A549
    suggested: None
    0.2 50 0.1 0 0.0 ay D ay D 3000 3000 1000 0 0 4 8 12 16 20 24 Time h 2000 1000 0 100 4ºC RT 0 100 200 300 400 Time, h % of total NCM 4000 Peak area 4000 2000 supernatant pellet E D PDI 0.3 100 5 tc h Ba tc h Ba tc h Ba tc h tc h Ba Ba C Peak area 0.4 0 0.0 4 0 3 0.1 2 50 150 42 0.2 0.5 7 0.3 100 200 ay 150 0.4 D 0.5 Diamter, nm 200 PDI B 1 Diamter, nm A 80 60 40 20 0 1 4 24 Time h Fig. 3. NCM IC50 0.042µM IC90 0.251µM FFU/mL (Log10) 8 7 6 5 4 3 2 -1.0 -0.5 0.0 0.5 1.0 NCM Concentration uM (Log10) -0.5 0.0 0.5 1.0 100 50 0 -1 0 1 2 NCM Concentration uM (Log10) nano NCM CC50 31.3µM 150 100 50 0 -1 0 1 2 NCM Concentration (uM) hACE2-A549 - nano NCM Nano NCM IC50 0.154µM; 8 IC90 1.38µM 7 6 5 4 3 2 -1.0 -0.5 0.0 0.5 1.0 NCM Concentration uM (Log10) hACE2-A549 viability, % FFU/mL (Log10) Vero cell viability, % 9 8 7 6 5 4 3 2 -1.0 nano NCM IC50 0.595µM IC90 3.38µM NCM Concentration uM (Log10) C NCM CC50 19.5µM 150 Vero E6 - nano NCM FFU/mL (Log10) B Vero cell viability, % Vero E6 - NCM A 150 Nano NCM CC50 21.13µM 100 50 0 -1 0 1 2 NCM Concentration uM (Log10) Fig.
    Vero
    suggested: None
    Vero E6
    suggested: None
    Software and Algorithms
    SentencesResources
    The IC50 and CC50 were determined by fitting normalized data to variable inhibition slope using Prism 8.0 software.
    Prism
    suggested: (PRISM, RRID:SCR_005375)
    Drug assay with HPLC - Agilent 1100 series equipped with Kinetex® 2.6 µm C18 100 Å, LC column 100 x 3 mm (Phenomenex Corporation, USA) was used.
    Kinetex®
    suggested: None
    Calibration curves for NCM were constructed using Agilent Masshunter Quantitative Analysis software.
    Agilent Masshunter Quantitative Analysis
    suggested: (Agilent Masshunter Quantitative Analysis software, RRID:SCR_015040)

    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: We found the following clinical trial numbers in your paper:

    IdentifierStatusTitle
    NCT03123978RecruitingEnzalutamide and Niclosamide in Treating Patients With Recur…
    NCT02807805RecruitingAbiraterone Acetate, Niclosamide, and Prednisone in Treating…
    NCT04436458Not yet recruitingNiclosamide In Moderate COVID-19
    NCT04399356RecruitingNiclosamide for Mild to Moderate COVID-19

    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.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  3. Version published to 10.1101/2020.12.18.423509v1 on bioRxiv