Multiomics approach reveals the ubiquitination-specific processes hijacked by SARS-CoV-2.

The Coronavirus Disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a global pandemic that seriously threatens health and socioeconomic development, but the existed antiviral drugs and vaccines still cannot yet halt the spread of the epidemic. Therefore, a comprehensive and profound understanding of the pathogenesis of SARS-CoV-2 is urgently needed to explore effective therapeutic targets. Here, we conducted a multiomics study of SARS-CoV-2-infected lung epithelial cells, including transcriptomic, proteomic, and ubiquitinomic. Multiomics analysis showed that SARS-CoV-2-infected lung epithelial cells activated strong innate immune response, including interferon and inflammatory responses. Ubiquitinomic further reveals the underlying mechanism of SARS-CoV-2 disrupting the host innate immune response. In addition, SARS-CoV-2 proteins were found to be ubiquitinated during infection despite the fact that SARS-CoV-2 itself didn't code any E3 ligase, and that ubiquitination at three sites on the Spike protein could significantly enhance viral infection. Further screening of the E3 ubiquitin ligases and deubiquitinating enzymes (DUBs) library revealed four E3 ligases influencing SARS-CoV-2 infection, thus providing several new antiviral targets. This multiomics combined with high-throughput screening study reveals that SARS-CoV-2 not only modulates innate immunity, but also promotes viral infection, by hijacking ubiquitination-specific processes, highlighting potential antiviral and anti-inflammation targets.

Antiviral efficacy of selective estrogen receptor modulators against SARS-CoV-2 infection in vitro and in vivo reveals bazedoxifene acetate as an entry inhibitor.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the seventh member of the coronavirus family that can infect humans. Recently, more contagious and pathogenic variants of SARS-CoV-2 have been continuously emerging. Clinical candidates with high efficacy and ready availability are still in urgent need. To identify potent anti-SARS-CoV-2 repurposing drugs, we evaluated the antiviral efficacy of 18 selective estrogen receptor modulators (SERMs) against SARS-CoV-2 infection. Six SERMs exhibited excellent anti-SARS-CoV-2 effects in Vero E6 cells and three human cell lines. Clomifene citrate, tamoxifen, toremifene citrate, and bazedoxifene acetate reduced the weight loss of hamsters challenged with SARS-CoV-2, and reduced hamster pulmonary viral load and interleukin-6 expression when assayed at 4 days postinfection. In particular, bazedoxifene acetate was identified to act on the penetration stage of the postattachment step via altering cholesterol distribution and endosome acidification. And, bazedoxifene acetate inhibited pseudoviruses infection of original SARS-CoV-2, Delta variant, Omicron variant, and SARS-CoV. These results offer critical information supporting bazedoxifene acetate as a promising agent against coronaviruses.

Discovery of potential anti-SARS-CoV-2 drugs based on large-scale screening in vitro and effect evaluation in vivo.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a global crisis. Clinical candidates with high efficacy, ready availability, and that do not develop resistance are in urgent need. Despite that screening to repurpose clinically approved drugs has provided a variety of hits shown to be effective against SARS-CoV-2 infection in cell culture, there are few confirmed antiviral candidates in vivo. In this study, 94 compounds showing high antiviral activity against SARS-CoV-2 in Vero E6 cells were identified from 2,580 FDA-approved small-molecule drugs. Among them, 24 compounds with low cytotoxicity were selected, and of these, 17 compounds also effectively suppressed SARS-CoV-2 infection in HeLa cells transduced with human ACE2. Six compounds disturb multiple processes of the SARS-CoV-2 life cycle. Their prophylactic efficacies were determined in vivo using Syrian hamsters challenged with SARS-CoV-2 infection. Seven compounds reduced weight loss and promoted weight regain of hamsters infected not only with the original strain but also the D614G variant. Except for cisatracurium, six compounds reduced hamster pulmonary viral load, and IL-6 and TNF-α mRNA when assayed at 4 d postinfection. In particular, sertraline, salinomycin, and gilteritinib showed similar protective effects as remdesivir in vivo and did not induce antiviral drug resistance after 10 serial passages of SARS-CoV-2 in vitro, suggesting promising application for COVID-19 treatment.

Isolation and identification of two strains of severe acute respiratory syndrome coronavirus 2 from coronavirus disease 2019 patients in Shanghai

Objective To isolate the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from nasal/throat swabs of coronavirus disease 19 (COVID-19) patients. Methods Three nasal/throat swab samples from COVID-19 patients in Shanghai were treated with TPCK trypsin and were used to treat Vero E6 cells inoculated in 96-well plates. When most of the cells showed obvious cytopathy, the cell culture supernatants were collected. We then detected the viral nucleic acid by fluorescent quantitative real-time polymerase chain reaction, and amplified the gene fragment of the virus receptor binding domain (RBD) by reverse transcription polymerase chain reaction. After amplification and culture, the virus was used to infect the Vero E6 cells inoculated in 96-well plates. The cytopathy was observed and the virus protein was detected by immunofluorescence. Results The Vero E6 cells that cultured with two of three nasal/pharyngeal swab samples showed obvious cytopathic effect and newly synthesized viral nucleic acid was detected in the supernatants of the cell culture. The amplified RBD sequence was completely consistent with the corresponding fragment of SARS-CoV-2 isolated earlier. Virus-infected Vero E6 cells showed cytopathies rapidly and could react with the monoclonal antibody against nucleocapsid protein (N protein) and spike protein (S protein) of SARS-CoV-2, and convalescence sera of COVID-19 patients. Conclusion Two SARS-CoV-2 strains were successfully isolated from two nasal/throat swab samples of COVID-19 patients in Shanghai, which provides evidence for the mechanism research on the infection and pathogenesis of SARS-CoV-2 as well as the development of drugs and vaccines against SARS-CoV-2.

Preparation and verification of severe acute respiratory syndrome Coronavirus 2 Pseudoparticles

Objective: To establish a method for preparing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudoparticles (SARS-CoV-2 pps).