Control of human coronavirus replication and virus-induced inflammatory processes by the NSAID indomethacin

Coronaviruses (CoV), zoonotic viruses periodically emerging worldwide, represent a constant potential threat to humans. To date, seven human coronaviruses (HCoV) have been identified: four endemic seasonal HCoV (sHCoV) and three highly-pathogenic HCoV (HP-HCoV). Although sHCoV generally cause only mild respiratory diseases in immunocompetent hosts, severe complications may occur in specific populations. Currently, there is no specific treatment for sHCoV infections. We now show that the nonsteroidal anti-inflammatory drug (NSAID) indomethacin (INDO), a potent cyclooxygenase (COX-1/COX-2) inhibitor, has remarkable cytoprotective and antiviral activity against seasonal coronaviruses α-HCoV-229E and βHCoV-OC43 in human lung cells. INDO does not affect sHCoV adsorption, entry or uncoating, but acts at post-entry level, causing a dose-dependent decrease in viral RNA and structural proteins expression. This effect is cyclooxygenase-independent and is associated with transient phosphorylation of eukaryotic initiation factor eIF2α at Ser51; besides, INDO antiviral activity is impaired in PKR-/- cells, identifying the eIF2α-kinase PKR as a molecular target for INDO in HCoV infection. Notably, in transfection experiments, INDO was also found to interfere with SARSCoV-2 spike glycoprotein stability. Moreover, we show that HCoV-229E and HCoV-OC43 infection provokes a high increase in COX-2 expression, triggering of a pro-inflammatory pathway in human lung cells; this event may be a direct consequence of the recently described activation of the transcription factor HSF1 during sHCoV infection. Interestingly, INDO, in addition to inhibiting sHCoV replication, also decreases sHCoV-induced COX-2 expression in human cells. More importantly, INDO inhibits HCoVinduced HSF1-Ser326 phosphorylation and transcriptional activation, which is essential for efficient HCoV replication, suggesting that inhibition of HSF1- signaling may contribute to INDO anti-CoV activity. Finally, because of the recent surge in SARS-CoV-2 and hRSV (human Respiratory Syncytial Virus) co-infections, the effect of INDO was investigated in hRSV-infected cells. Similarly to HCoV infection, we demonstrate that INDO potently inhibits hRSV replication in human cells, acting at post-entry level. Altogether the results suggest that indomethacin, due to the broad-spectrum antiviral activity combined with anti-inflammatory action, could represent a first-line of defense against both HCoV and hRSV infections.