Cell cycle block by p53 activation reduces SARS-CoV-2 release in infected alveolar basal epithelial A549-hACE2 cells

Abstract

SARS-CoV viruses have been shown to downregulate cellular events that control antiviral defenses. They adopt several strategies to silence p53, key molecule for cell homeostasis and immune control, indicating that p53 has a central role in controlling their proliferation in the host. Specific actions are the stabilization of its inhibitor, MDM2, and the interference with its transcriptional activity. The aim of our work was to evaluate a new approach against SARS-CoV-2 by using MDM2 inhibitors to raise p53 levels and activate p53-dependent pathways, therefore leading to cell cycle inhibition. Experimental setting was performed in the alveolar basal epithelial cell line A549-hACE2, expressing high level of ACE2 receptor, to allow virus entry, as well as p53 wild-type. Cells were treated with several concentrations of Nutlin-3 or RG-7112, two known MDM2 inhibitors, for the instauration of a cell cycle block steady-state condition before and during SARS-CoV-2 infection, and for the evaluation of p53 activation and impact on virus release and related innate immune events. The results indicated an efficient cell cycle block with inhibition of the virion release and a significant inhibition of IL-6, NF-kB and IFN-λ expression. These data suggest that p53 is an efficient target for new therapies against the virus and that MDM2 inhibitors deserve to be further investigated in this field.

P53-Independent G1-Cell Cycle Arrest Increases SARS-CoV-2 RNA Replication

“…Several studies have previously highlighted the role of p53 in SARS-CoV-2 infection [ 34 , 35 ], pointing out p53 as a potential target for the development of antivirals [ 36 , 37 ]. In contrast, our data show that the increase in SARS-CoV-2 replication observed when the cells are stopped in G1 is p53-independent, making the use of p53 targeting antivirals ineffective in this case.…”

Cell-Dependent Activation of ProTide Prodrugs and Its Implications in Antiviral Studies

“…Vero E6, A549, Calu-3, Caco-2, and Huh-7 are among the most commonly used cell lines for evaluating the antiviral activity of antiviral drugs in virology studies . Among these, Vero E6 cells have been widely used as an in vitro cell model for anti-SARS-CoV-2 research due to its outstanding efficiency in propagating SARS-CoV-2. , The lung cell lines A459 and Calu-3 are commonly used as in vitro pneumonocyte models for the research of pulmonary infections such as COVID-19. − The human colon cell line Caco-2 and human liver cell line Huh-7 have been adopted as in vitro models for the study of colon and hepatitis virus infections. , The antiviral activity of a prodrug depends on the intracellular accumulation and activation of the prodrug. Previous in vitro studies have shown that the activation and antiviral efficacy of nucleoside/nucleotide prodrugs, such as remdesivir, were cell-line-dependent. , Thus, the in vitro antiviral activity results could be misleading when the prodrug is not adequately activated in the selected cells.…”

State of the Art of Pharmacological Activators of p53 in Ocular Malignancies

“…Among these, Nutlin-3 is a synthetic small molecule cis-imidazoline analog that mimics highly conserved hydrophobic amino acid residues including Phe19, Trp23, Leu22, and Leu26 within the hydrophobic pocket of MDM2 [126]. Nutlins have been shown to induce cell cycle arrest and cell death in a variety of solid tumors as well as in several types of hematological malignancies, viral infections, and cancer models with wild-type p53 including osteosarcoma, prostate cancer, Kaposi's sarcoma-associated herpesvirus lymphomas, and neuroblastoma [128][129][130][131][132][133][134].…”