Discovery of re-purposed drugs that slow SARS-CoV-2 replication in human cells

Abstract

Background: The SARS-CoV-2 virus has caused the death of over 2 million people worldwide during the COVID-19 pandemic. Whilst effective vaccines have been developed and vaccination schedules are being rolled out, the identification of safe and inexpensive drugs to slow the replication of SARS-CoV-2 could help thousands of people worldwide whilst awaiting vaccination. Methods: Using SARS-CoV-2 tagged with nano-luciferase (SARS-CoV-2-ΔOrf7a-NLuc) we screened a variety of cells under optimised cell culture conditions for their ability to be infected by, and support the replication of, SARS-CoV-2. Electron microscopy was used to demonstrate generation of infectious virus particles. We assessed a library of 1971 FDA-approved drugs for their ability to inhibit or enhance viral replication in Vero (simian kidney cells) but also in the human hepatocyte cell, HUH7. Initial hits were further tested to identify compounds that could suppress viral replication, post-viral infection. Dose response curves were obtained for a shortlist of 9 compounds of interest (COI). Findings: Our SARS-CoV-2-ΔOrf7a-NLuc virus was as effective as wild-type SARS-CoV-2 in inducing CPE and replicating in Vero cells. Conventional electron microscopy showed the NLuc-tagged virus to be structurally indistinguishable from the wild-type virus, and both could be identified within the endosomal system of infected cells. SARS-CoV-2-ΔOrf7a-NLuc was used in experiments to robustly quantitate virus infection and replication. A wide variety of human cells including lung fibroblasts and epithelial cells were susceptible to infection but were not effective in supporting SARS-CoV-2-ΔOrf7a-NLuc replication. In contrast, human kidney epithelial cells and human hepatic cells were particularly susceptible and supported SARS-CoV-2-replication, which is in-line with reported proteinuria and liver damage in patients with COVID-19. Our screening of FDA approved compounds identified 35 COI that inhibited virus infection and replication in either Vero or human cell lines. Nine of these also inhibited SARS-CoV-2 replication when treatment commenced after virus infection. Therapeutics approved for treatment of cancer, malaria, hypertension and viral infection were identified with atovaquone, manidipine, vitamin D3 and ebastine being well tolerated with minimal side effects. Only two COI were consistently found to enhance SARS-CoV-2 replication, aliskiren and lithocholic acid. Interpretation: Re-purposing of safe, well-tolerated FDA-approved drugs that inhibit SARS-CoV-2 replication is an attractive strategy to reduce the risk of COVID-19 infection prior to receiving an effective vaccine. The COI identified here hold potential to contain COVID-19 whilst wide-scale vaccination proceeds. The identification of FDA-approved drugs that enhance SARS-CoV-2 replication in human cells suggests that entry routes into cells can be made more accessible to the virus by certain medications. The information provided in this research paper is for information only and is not meant to be a substitute for advice provided by a doctor or other qualified health care professional.

Computational screening of potential drugs against COVID-19 disease: the Neuropilin-1 receptor as molecular target

“…In vitro screening of 30 approved medicines reveals that Ebastine displayed a strong inhibition of the SARS-CoV-2 main protease (Mpro) in infected cells [24]. In addition, a preprint paper reported the ability of this in slowing the replication process of SARS-CoV-2 in human cells [17].…”

Mutual inhibition of airway epithelial responses supports viral and fungal co-pathogenesis during coinfection

“…We previously reported that SARS-CoV2 was able to infect A549 epithelial cells., but showed limited replicative capacity. Increased NLuc signal, indicating viral replication (29), was detected when monolayers were exposed to Af culture filtrates from Af293 but not A1160 strains (P < 0.05) (Figure 2c) . Interestingly, heightened NLuc activity was detected when A549 cells were co-cultured with THP1 monocytes and Af293 culture filtrates (Supplementary Figure 2b) .…”

“…Equivalent quantities were used in the staining mix and fluorescence for this dye was acquired in Channel 5 of the IFC. Live-cell imaging and analyses of fungal uptake and intracellular killing were performed as described in (13) (29). NLuc activity was measured at 72 h post-infection in the presence of 0.5 L of coelenterazine as previously reported (29).…”

“…Live-cell imaging and analyses of fungal uptake and intracellular killing were performed as described in (13) (29). NLuc activity was measured at 72 h post-infection in the presence of 0.5 L of coelenterazine as previously reported (29). Statistical analyses were done using Prism GraphPad v9 (La Joya, CA).…”

“…On the basis of these observations we further hypothesised that secreted factors produced by Af during vegetative growth are likely regulators of viral pathogenicity. To model viral-fungal coinfection during fungal germination, A549 monolayers were exposed to live SARS-CoV expressing a Nanoluciferase reporter (SARS-CoV-2-ΔOrf7a-NLuc (29)) in the presence or absence of Af A1160 and Af293 culture filtrates recapitulating secreted factors during hyphal growth. These two Af reference strains were chosen as they have different protease activity (30, 31).…”