SARS-CoV-2 Fears Green: The Chlorophyll Catabolite Pheophorbide A Is a Potent Antiviral

  1. Guillermo H. Jimenez-Aleman
  2. Victoria Castro
  3. Addis Londaitsbehere
  4. Marta Gutierrez-Rodríguez
  5. Urtzi Garaigorta
  6. Roberto Solano
  7. Pablo Gastaminza

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Abstract

SARS-CoV-2 pandemic is having devastating consequences worldwide. Although vaccination advances at good pace, effectiveness against emerging variants is unpredictable. The virus has displayed a remarkable resistance to treatments and no drugs have been proved fully effective against COVID-19. Thus, despite the international efforts, there is still an urgent need for new potent and safe antivirals against SARS-CoV-2. Here, we exploited the enormous potential of plant metabolism using the bryophyte Marchantia polymorpha L. and identified a potent SARS-CoV-2 antiviral, following a bioactivity-guided fractionation and mass-spectrometry approach. We found that the chlorophyll derivative Pheophorbide a (PheoA), a porphyrin compound similar to animal Protoporphyrin IX, has an extraordinary antiviral activity against SARS-CoV-2, preventing infection of cultured monkey and human cells, without noticeable cytotoxicity. We also show that PheoA targets the viral particle, interfering with its infectivity in a dose- and time-dependent manner. Besides SARS-CoV-2, PheoA also displayed a broad-spectrum antiviral activity against enveloped RNA viral pathogens such as HCV, West Nile, and other coronaviruses. Our results indicate that PheoA displays a remarkable potency and a satisfactory therapeutic index, which together with its previous use in photoactivable cancer therapy in humans, suggest that it may be considered as a potential candidate for antiviral therapy against SARS-CoV-2.

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  1. Version published to 10.3390/ph14101048
  2. Version published to 10.1101/2021.07.31.454592v2 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2021.07.31.454592: (What is this?)

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

    Table 1: Rigor

    Ethicsnot detected.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    A monoclonal antibody against the N protein was diluted in the incubation buffer (1:2000, v/v; Genetex HL344) and incubated with the cells for 1 h; after this time, cells were washed with PBS and subsequently incubated with a 1:500 (v/v) dilution of a goat anti-rabbit conjugated to Alexa 488 (Invitrogen-Carlsbad, CA).
    anti-rabbit
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Cell culture: Vero E6 (ATCC) and Calu3 (ATCC) cell lines were kindly provided by Dr. Enjuanes (CNB-CSIC)
    Calu3
    suggested: BCRJ Cat# 0264, RRID:CVCL_0609)
    A549 and Huh7 cells were transduced with a retroviral vector enabling expression of ACE2 in a di-cistronic expression cassette also conferring resistance to blasticidine.
    A549
    suggested: NCI-DTP Cat# A549, RRID:CVCL_0023)
    The hCoV-229E-GFP51 was kindly provided by Dr. Thiel (University of Basel) and propagated in Huh7 cells at 33°C in a controlled 5% CO2 environment.
    Huh7
    suggested: None
    Cypopathic effect protection assays in Vero E6 and Huh7-ACE2 cells: Vero E6 or Huh7-ACE2 cell monolayers were inoculated (MOI = 0.001) in the presence of a wide range of two-fold dilutions of the crude, or partially purified extracts, or pure compounds and incubated for 72 h.
    Vero E6
    suggested: None
    Huh7-ACE2
    suggested: None
    Assessment of viral entry using retroviral pseudotypes: Retroviral particles pseudotyped with SARS-2-CoV spike envelope protein (Spp) were produced in HEK293T cells as previously described54 with materials kindly provided by Dr. F. L. Cosset (INSERM, Lyon) and J. M. Casasnovas and J. G. Arriaza (CNB-CSIC) for the S protein cDNA.
    HEK293T
    suggested: None
    Recombinant DNA
    SentencesResources
    VSV-GFP30 was kindly provided by Dr. Rodriguez (CNB-CSIC).
    VSV-GFP30
    suggested: None
    Software and Algorithms
    SentencesResources
    Statitistical Analysis: Descriptive statistics were calculated using Microsoft Excel.
    Microsoft Excel
    suggested: (Microsoft Excel, RRID:SCR_016137)
    One-way ANOVA and post-hoc tests were calculated using IBM SPSS Software Package (version 26).
    SPSS
    suggested: (SPSS, RRID:SCR_002865)

    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.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

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  4. Version published to 10.1101/2021.07.31.454592v1 on bioRxiv