Project description:The SARS-CoV-2 pandemic, and the likelihood of future coronavirus pandemics, has rendered our understanding of coronavirus biology more essential than ever. Small molecule chemical probes offer to both reveal novel aspects of virus replication and to serve as leads for antiviral therapeutic development. The RNA-biased amiloride scaffold was recently tuned to target a viral RNA structure critical for translation in enterovirus 71, ultimately uncovering a novel mechanism to modulate positive-sense RNA viral translation and replication. Analysis of CoV RNA genomes reveal many conserved RNA structures in the 5'-UTR and proximal region critical for viral translation and replication, including several containing bulge-like secondary structures suitable for small molecule targeting. Following phylogenetic conservation analysis of this region, we screened an amiloride-based small molecule library against a less virulent human coronavirus, OC43, to identify lead ligands. Amilorides inhibited OC43 replication as seen in viral plaque assays. Select amilorides also potently inhibited replication competent SARS-CoV-2 as evident in the decreased levels of cell free virions in cell culture supernatants of treated cells. Reporter screens confirmed the importance of RNA structures in the 5'-end of the viral genome for small molecule activity. Finally, NMR chemical shift perturbation studies of the first six stem loops of the 5'-end revealed specific amiloride interactions with stem loops 4, 5a, and 6, all of which contain bulge like structures and were predicted to be strongly bound by the lead amilorides in retrospective docking studies. Taken together, the use of multiple orthogonal approaches allowed us to identify the first small molecules aimed at targeting RNA structures within the 5'-UTR and proximal region of the CoV genome. These molecules will serve as chemical probes to further understand CoV RNA biology and can pave the way for the development of specific CoV RNA-targeted antivirals.
Project description:An escalating pandemic by the novel SARS-CoV-2 virus is impacting global health and effective therapeutic options are urgently needed. We evaluated the in vitro antiviral effect of compounds that were previously reported to inhibit coronavirus replication and compounds that are currently under evaluation in clinical trials for SARS-CoV-2 patients. We report the antiviral effect of remdesivir, lopinavir, homorringtonine, and emetine against SARS-CoV-2 virus in Vero E6 cells with the estimated 50% effective concentration at 23.15 ?M, 26.63 ?M, 2.55 ?M and 0.46 ?M, respectively. Ribavirin or favipiravir that are currently evaluated under clinical trials showed no inhibition at 100 ?M. Synergy between remdesivir and emetine was observed, and remdesivir at 6.25 ?M in combination with emetine at 0.195 ?M may achieve 64.9% inhibition in viral yield. Combinational therapy may help to reduce the effective concentration of compounds below the therapeutic plasma concentrations and provide better clinical benefits.
Project description:In December 2019, a new severe acute respiratory syndrome coronavirus (SARS-CoV-2) causing coronavirus diseases 2019 (COVID-19) emerged in Wuhan, China. African countries see slower dynamic of COVID-19 cases and deaths. One of the assumptions that may explain this later emergence in Africa, and more particularly in malaria endemic areas, would be the use of antimalarial drugs. We investigated the in vitro antiviral activity against SARS-CoV-2 of several antimalarial drugs. Chloroquine (EC50 = 2.1 μM and EC90 = 3.8 μM), hydroxychloroquine (EC50 = 1.5 μM and EC90 = 3.0 μM), ferroquine (EC50 = 1.5 μM and EC90 = 2.4 μM), desethylamodiaquine (EC50 = 0.52 μM and EC90 = 1.9 μM), mefloquine (EC50 = 1.8 μM and EC90 = 8.1 μM), pyronaridine (EC50 = 0.72 μM and EC90 = 0.75 μM) and quinine (EC50 = 10.7 μM and EC90 = 38.8 μM) showed in vitro antiviral effective activity with IC50 and IC90 compatible with drug oral uptake at doses commonly administered in malaria treatment. The ratio Clung/EC90 ranged from 5 to 59. Lumefantrine, piperaquine and dihydroartemisinin had IC50 and IC90 too high to be compatible with expected plasma concentrations (ratio Cmax/EC90 < 0.05). Based on our results, we would expect that countries which commonly use artesunate-amodiaquine or artesunate-mefloquine report fewer cases and deaths than those using artemether-lumefantrine or dihydroartemisinin-piperaquine. It could be necessary now to compare the antimalarial use and the dynamics of COVID-19 country by country to confirm this hypothesis.
Project description:The untranslated regions (UTRs) of viral genomes contain a variety of conserved yet dynamic structures crucial for viral replication, providing drug targets for the development of broad spectrum anti-virals. We combine in vitro RNA analysis with molecular dynamics simulations to build the first 3D models of the structure and dynamics of key regions of the 5' UTR of the SARS-CoV-2 genome. Furthermore, we determine the binding of metallo-supramolecular helicates (cylinders) to this RNA structure. These nano-size agents are uniquely able to thread through RNA junctions and we identify their binding to a 3-base bulge and the central cross 4-way junction located in stem loop 5. Finally, we show these RNA-binding cylinders suppress SARS-CoV-2 replication, highlighting their potential as novel anti-viral agents.
Project description:Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has become one major threat to human population health. The RNA-dependent RNA polymerase (RdRp) presents an ideal target of antivirals, whereas nucleoside analogs inhibitor is hindered by the proofreading activity of coronavirus. Herein, we report that corilagin (RAI-S-37) as a non-nucleoside inhibitor of SARS-CoV-2 RdRp, binds directly to RdRp, effectively inhibits the polymerase activity in both cell-free and cell-based assays, fully resists the proofreading activity and potently inhibits SARS-CoV-2 infection with a low 50% effective concentration (EC50) value of 0.13 μmol/L. Computation modeling predicts that RAI-S-37 lands at the palm domain of RdRp and prevents conformational changes required for nucleotide incorporation by RdRp. In addition, combination of RAI-S-37 with remdesivir exhibits additive activity against anti-SARS-CoV-2 RdRp. Together with the current data available on the safety and pharmacokinetics of corilagin as a medicinal herbal agent, these results demonstrate the potential of being developed into one of the much-needed SARS-CoV-2 therapeutics.
Project description:Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged as a new human pathogen in late 2019 and it has infected over 100 million people in less than a year. There is a clear need for effective antiviral drugs to complement current preventive measures, including vaccines. In this study, we demonstrate that berberine and obatoclax, two broad-spectrum antiviral compounds, are effective against multiple isolates of SARS-CoV-2. Berberine, a plant-derived alkaloid, inhibited SARS-CoV-2 at low micromolar concentrations and obatoclax, which was originally developed as an anti-apoptotic protein antagonist, was effective at sub-micromolar concentrations. Time-of-addition studies indicated that berberine acts on the late stage of the viral life cycle. In agreement, berberine mildly affected viral RNA synthesis, but it strongly reduced infectious viral titers, leading to an increase in the particle-to-pfu ratio. In contrast, obatoclax acted at the early stage of the infection, which is in line with its activity to neutralize the acidic environment in endosomes. We assessed infection of primary human nasal epithelial cells that were cultured on an air-liquid interface and found that SARS-CoV-2 infection induced and repressed expression of specific sets of cytokines and chemokines. Moreover, both obatoclax and berberine inhibited SARS-CoV-2 replication in these primary target cells. We propose berberine and obatoclax as potential antiviral drugs against SARS-CoV-2 that could be considered for further efficacy testing.
Project description:The ongoing COVID-19/Severe Acute Respiratory Syndrome CoV-2 (SARS-CoV-2) pandemic has become a significant threat to public health and has hugely impacted societies globally. Targeting conserved SARS-CoV-2 RNA structures and sequences essential for viral genome translation is a novel approach to inhibit viral infection and progression. This new pharmacological modality compasses two classes of RNA-targeting molecules: 1) synthetic small molecules that recognize secondary or tertiary RNA structures and 2) antisense oligonucleotides (ASOs) that recognize the RNA primary sequence. These molecules can also serve as a "bait" fragment in RNA degrading chimeras to eliminate the viral RNA genome. This new type of chimeric RNA degrader is recently named ribonuclease targeting chimera or RIBOTAC. This review paper summarizes the sequence conservation in SARS-CoV-2 and the current development of RNA-targeting molecules to combat this virus. These RNA-binding molecules will also serve as an emerging class of antiviral drug candidates that might pivot to address future viral outbreaks.
Project description:Translation of open reading frame 1b (ORF1b) in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) requires programmed -1 ribosomal frameshifting (-1 PRF) promoted by an RNA pseudoknot. The extent to which SARS-CoV-2 replication may be sensitive to changes in -1 PRF efficiency is currently unknown. Through an unbiased, reporter-based high-throughput compound screen, we identified merafloxacin, a fluoroquinolone antibacterial, as a -1 PRF inhibitor of SARS-CoV-2. Frameshift inhibition by merafloxacin is robust to mutations within the pseudoknot region and is similarly effective on -1 PRF of other beta coronaviruses. Importantly, frameshift inhibition by merafloxacin substantially impedes SARS-CoV-2 replication in Vero E6 cells, thereby providing the proof of principle of targeting -1 PRF as an effective antiviral strategy for SARS-CoV-2.