A robust SARS-CoV-2 replication model in primary human epithelial cells at the air liquid interface to assess antiviral agents

Abstract

There are, besides Remdesivir (RDV), no approved antivirals for the treatment and/or prophylaxis of SARS-CoV-2 infections. To aid in the search for antivirals against this virus, we explored the use of human tracheal airway epithelial cells (HAEC) and human small airway epithelial cells (HsAEC) grown at the air/liquid interface (ALI) and infected at the apical side with either one of two different SARS-CoV-2 isolates. The virus was shown to replicate to high titers for extended periods of time (at least 8 days) and, in particular an isolate with the D614G in the spike (S) protein did so more efficiently at 35°C than at 37°C. The effect of a selected panel of reference drugs that were added to the culture medium at the basolateral side of the system was explored. GS-441524 (the parent nucleoside of Remdesivir), EIDD-1931 (the active metabolite of Molnupiravir) and IFN (β1 and λ1) all resulted in a dose-dependent inhibition of viral RNA and infectious virus titers at the apical side. However, AT-511 (a guanosine nucleotide previously reported to inhibit SARS-CoV-2) failed to inhibit viral replication. Together, these results provide a reference for further studies aimed at selecting SARS-CoV-2 inhibitors for further preclinical and clinical development.

Comparison of anti-SARS-CoV-2 activity and intracellular metabolism of remdesivir and its parent nucleoside

“…Finally, primary HAE cells have been used widely by researchers to study the SARS-CoV-2 ( Sheahan et al, 2020 ; Pizzorno et al, 2020 ; Hattori et al, 2021 ; Mulay et al, 2021 ; Do et al, 2021 ). Researchers compared the anti-SARS-CoV-2 activity of remdesivir and GS-441524 in HAE models and showed that remdesivir was more potent than GS-441524 in human tracheal airway cultures ( Do et al, 2021 ). In HAE cells at 4 h, remdesivir generated about 20-fold more (p < 0.001, t -test) GS-443902 than the parent nucleoside GS-441524 ( Fig.…”

Remdesivir and GS-441524 retain antiviral activity against Delta, Omicron, and other emergent SARS-CoV-2 variants

“…Since the beginning of pandemic, WHO declared 5 of these variants as VOCs that could be associated with more severe disease and/or increased rate of transmission. While the SARS-CoV-2 antiviral activity of RDV has previously been well characterized both in vitro and in vivo (13,24,26,(29)(30)(31), most studies have been conducted using the ancestral WA1 isolate. Here, we sought to fully characterize the antiviral potency of RDV and its parent nucleoside GS-441524 against a panel of the most significant SARS-CoV-2 variants including all the major VOCs.…”

“…The RdRp catalytic active site is nearly 100% conserved among coronaviruses, therefore the observed potency of RDV against other coronaviruses was anticipated to translate to SARS-CoV-2 antiviral activity (21). RDV and GS-441524 have both demonstrated potency against SARS-CoV-2, with in vitro cellular EC50 values ranging from 10 to 120 nM for RDV and 470 to 3600 nM for GS-441524 (13,(24)(25)(26)(27). The in vivo efficacy of RDV has been demonstrated in SARS-CoV-2 challenge studies in mice and hamsters (13,28,29).…”

Mechanisms of Lung Injury Induced by SARS-CoV-2 Infection

“…An early report used human primary airway epithelial cells cultured on air-liquid phase to validate the viral infection and cytopathic effect ( 62 ). The human primary tracheal airway epithelial cells and small airway epithelial cells were further used to evaluate antiviral drug candidates ( 63 ). However, primary lung epithelial cells can be challenging to expand in culture and also exhibit significant patient-to-patient or batch-to-batch variability.…”