Potential antiviral effects of pantethine against SARS-CoV-2.

SARS-CoV-2 interacts with cellular cholesterol during many stages of its replication cycle. Pantethine was reported to reduce total cholesterol levels and fatty acid synthesis and potentially alter different processes that might be involved in the SARS-CoV-2 replication cycle. Here, we explored the potential antiviral effects of pantethine in two in vitro experimental models of SARS-CoV-2 infection, in Vero E6 cells and in Calu-3a cells. Pantethine reduced the infection of cells by SARS-CoV-2 in both preinfection and postinfection treatment regimens. Accordingly, cellular expression of the viral spike and nucleocapsid proteins was substantially reduced, and we observed a significant reduction in viral copy numbers in the supernatant of cells treated with pantethine. In addition, pantethine inhibited the infection-induced increase in TMPRSS2 and HECT E3 ligase expression in infected cells as well as the increase in antiviral interferon-beta response and inflammatory gene expression in Calu-3a cells. Our results demonstrate that pantethine, which is well tolerated in humans, was very effective in controlling SARS-CoV-2 infection and might represent a new therapeutic drug that can be repurposed for the prevention or treatment of COVID-19 and long COVID syndrome.

Bronchial epithelia from adults and children: SARS-CoV-2 spread via syncytia formation and Type-III interferon infectivity restriction

Bronchi of the upper respiratory tract are considered the site of SARSCoV- 2 infection initiation from where a possible spread to the lower respiratory tract can cause acute respiratory distress syndrome with a high degree of mortality in elderly patients. Here we established functional reconstituted primary bronchial epithelia (BE) derived from donors including both adults and children to study SARS-CoV-2 infection dynamics in a physiologically relevant model. We identified multi-ciliated cells as the primary target cells for SARS-CoV-2 in our reconstituted BE. We further observed rapid viral spread throughout the entire BE within 24-48hours. Within 3-4 days, we observed syncytia formation between ciliated cells and basal cells which accumulate at the apical side of the BE. We show that infected cells including syncytia are released into the apical lumen and contribute to the transmittable virus dose. Interestingly, some BE mainly reconstituted from young donor, showed an intrinsic resistance to infection and virus spread. This restricted infection phenotype correlated with a faster release of type-III interferon secretion. Moreover, exogenous type-III interferon treatment to permissive epithelia installed infection restriction while interferon gene knockout promoted infection. Taken together our data uncover syncytia formation as possible contribution to tissue or environmental SARS-CoV-2 dissemination and the type-III IFN response as a central contributor to SARS-CoV-2 resistance in BE, which may explain epidemiological observations that SARS-CoV-2 fatality is age dependent.

Rational development of tools to appropriately respond to SARSCoV- 2 needs

For many years, our laboratory has been developing cellular models for the study of human pathogenic viruses with RNA genomes, in order to study the replication of these pathogens, to propose new therapeutic pathways, to screen and test inhibitors. In response to the COVID-19 outbreak, we have set up the tools for the study of SARS-CoV-2 replication. First, clinical and reference SARS-CoV-2 strains have been successfully isolated and amplified using Vero E6 cells in the BSL3 facility of Bordeaux University (UB'L3, www.mfp.cnrs.fr/wp/larecherche/ andevir/ubl3/). We set up the monitoring of SARS-CoV-2 replication using conventional RT-qPCR quantification as well as evaluation of the cytopathic effect by microscopic observation or content analysis. Using VERO cells, we are now able to precisely titer viral supernatant (determination of the TCID50) and screen for potential antiviral molecule (determination of EC50 and CC50). We have developed a full-length Spike sequencing based on a Sanger approach1 as well as whole genome sequencing by nanopore technology, allowing the tracking of emerging variants. In parallel, we developed various other models to study SARS-CoV-2 replication including Calu-3 cells, modified human cells expressing Ace2 (e.g. 293T, U2OS) or even more complex cellular models (reconstituted human airway epithelium, vessels) according to the biological question to resolve. As an example, bronchial epithelia reconstituted from biopsies of adult or child donors were used to evaluate the inflammatory response upon SARS-CoV-2 infection in an age-dependent manner [2] (see poster G. Beucher). Similarly reconstituted blood vessels were used to study the impact of SARS-CoV- 2 infection on the vascular system and determine whether clinical observations (blood brain barrier damages, myocarditis) are due to direct infection of cells or indirect effects. Finally, we evaluate the efficacy of different chemical or physical processes for viral inactivation in air or on surfaces.