A Novel Glucocorticoid and Androgen Receptor Modulator Reduces Viral Entry and Innate Immune Inflammatory Responses in the Syrian Hamster Model of SARS-CoV-2 Infection

  1. Savannah M. Rocha
  2. Anna C. Fagre
  3. Amanda S. Latham
  4. Jason E. Cummings
  5. Tawfik A. Aboellail
  6. Philip Reigan
  7. Devin A. Aldaz
  8. Casey P. McDermott
  9. Katriana A. Popichak
  10. Rebekah C. Kading
  11. Tony Schountz
  12. Neil D. Theise
  13. Richard A. Slayden
  14. Ronald B. Tjalkens

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Abstract

Despite significant research efforts, treatment options for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remain limited. This is due in part to a lack of therapeutics that increase host defense to the virus. Replication of SARS-CoV-2 in lung tissue is associated with marked infiltration of macrophages and activation of innate immune inflammatory responses that amplify tissue injury. Antagonists of the androgen (AR) and glucocorticoid (GR) receptors have shown efficacy in models of COVID-19 and in clinical studies because the cell surface proteins required for viral entry, angiotensin converting enzyme 2 (ACE2) and the transmembrane protease, serine 2 (TMPRSS2), are transcriptionally regulated by these receptors. We postulated that the GR and AR modulator, PT150, would reduce infectivity of SARS-CoV-2 and prevent inflammatory lung injury in the Syrian golden hamster model of COVID-19 by down-regulating expression of critical genes regulated through these receptors. Animals were infected intranasally with 2.5 × 10 4 TCID 50 /ml equivalents of SARS-CoV-2 (strain 2019-nCoV/USA-WA1/2020) and PT150 was administered by oral gavage at 30 and 100 mg/Kg/day for a total of 7 days. Animals were examined at 3, 5 and 7 days post-infection (DPI) for lung histopathology, viral load and production of proteins regulating the progression of SARS-CoV-2 infection. Results indicated that oral administration of PT150 caused a dose-dependent decrease in replication of SARS-CoV-2 in lung, as well as in expression of ACE2 and TMPRSS2. Lung hypercellularity and infiltration of macrophages and CD4 + T-cells were dramatically decreased in PT150-treated animals, as was tissue damage and expression of IL-6. Molecular docking studies suggest that PT150 binds to the co-activator interface of the ligand-binding domain of both AR and GR, thereby acting as an allosteric modulator and transcriptional repressor of these receptors. Phylogenetic analysis of AR and GR revealed a high degree of sequence identity maintained across multiple species, including humans, suggesting that the mechanism of action and therapeutic efficacy observed in Syrian hamsters would likely be predictive of positive outcomes in patients. PT150 is therefore a strong candidate for further clinical development for the treatment of COVID-19 across variants of SARS-CoV-2.

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  1. Version published to 10.3389/fimmu.2022.811430
  2. ScreenIT

    SciScore for 10.1101/2021.02.20.432110: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIACUC: Animal procedures: All animal protocols were approved by the Institutional Animal Use Committee at.
    RandomizationQuantification of protein was performed by acquiring five randomized images encompassing the pseudostratified columnar epithelium around bronchi at 400x magnification (Olympus X-Apochromat air objective; N.A. 0.95) all from different lung lobes.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variableMale and female Syrian hamsters were used in studies at 8 weeks of age (N=60, Charles River Laboratory).
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Primary antibodies were diluted to their optimized dilutions in tris-buffered saline and incubated on the tissue for 1 hour/antibody: Rabbit SARS nucleocapsid protein (SARS-CoV-2; Rockland; 1:500), goat ionized calcium binding adaptor molecule 1 (IBA1; Abcam; 1:50), goat angiotensin converting enzyme 2 (ACE2;R&D Systems; 1:500), rabbit transmembrane serine protease 2 (TMPRSS2; Abcam; 1:500)
    SARS nucleocapsid protein
    suggested: (Novus Cat# NB100-56576, RRID:AB_838838)
    ionized calcium binding adaptor molecule 1
    suggested: (Wako Cat# 01-1874, RRID:AB_2314666)
    angiotensin converting enzyme 2
    suggested: (Aviva Systems Biology Cat# ARP53751_P050, RRID:AB_2045325)
    Experimental Models: Cell Lines
    SentencesResources
    Virus and cell culture procedures: SARS-CoV-2 strain 2019-nCoV/USA-WA1/2020 was obtained from BEI Resources and propagated on Vero cells (ATCC CCL-81, American Type Culture Collection) at 37°C with 5% CO2.
    Vero
    suggested: None
    Software and Algorithms
    SentencesResources
    ROIs were drawn around the lung sections and the co-localization function of Count and Measure within Olympus CellSens software was applied the sections.
    CellSens
    suggested: None
    PT150 prepared using LigPrep by generating possible states at the target pH 7.0 using Epik and minimized by applying the OPLS_2005 force field [27].
    LigPrep
    suggested: (Ligprep, RRID:SCR_016746)

    Results from OddPub: Thank you for sharing your data.

    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: Please consider improving the rainbow (“jet”) colormap(s) used on pages 33, 34, 35 and 36. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    About SciScore

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  3. Version published to 10.1101/2021.02.20.432110v1 on bioRxiv