Design of SARS-CoV-2 PLpro Inhibitors for COVID-19 Antiviral Therapy Leveraging Binding Cooperativity.

Antiviral agents that complement vaccination are urgently needed to end the COVID-19 pandemic. The SARS-CoV-2 papain-like protease (PLpro), one of only two essential cysteine proteases that regulate viral replication, also dysregulates host immune sensing by binding and deubiquitination of host protein substrates. PLpro is a promising therapeutic target, albeit challenging owing to featureless P1 and P2 sites recognizing glycine. To overcome this challenge, we leveraged the cooperativity of multiple shallow binding sites on the PLpro surface, yielding novel 2-phenylthiophenes with nanomolar inhibitory potency. New cocrystal structures confirmed that ligand binding induces new interactions with PLpro: by closing of the BL2 loop of PLpro forming a novel "BL2 groove" and by mimicking the binding interaction of ubiquitin with Glu167 of PLpro. Together, this binding cooperativity translates to the most potent PLpro inhibitors reported to date, with slow off-rates, improved binding affinities, and low micromolar antiviral potency in SARS-CoV-2-infected human cells.

Nomogram for Predicting COVID-19 Disease Progression Based on Single-Center Data: Observational Study and Model Development.

BACKGROUND: In late December 2019, a pneumonia caused by SARS-CoV-2 was first reported in Wuhan and spread worldwide rapidly. Currently, no specific medicine is available to treat infection with COVID-19. OBJECTIVE: The aims of this study were to summarize the epidemiological and clinical characteristics of 175 patients with SARS-CoV-2 infection who were hospitalized in Renmin Hospital of Wuhan University from January 1 to January 31, 2020, and to establish a tool to identify potential critical patients with COVID-19 and help clinical physicians prevent progression of this disease. METHODS: In this retrospective study, clinical characteristics of 175 confirmed COVID-19 cases were collected and analyzed. Univariate analysis and least absolute shrinkage and selection operator (LASSO) regression were used to select variables. Multivariate analysis was applied to identify independent risk factors in COVID-19 progression. We established a nomogram to evaluate the probability of progression of the condition of a patient with COVID-19 to severe within three weeks of disease onset. The nomogram was verified using calibration curves and receiver operating characteristic curves. RESULTS: A total of 18 variables were considered to be risk factors after the univariate regression analysis of the laboratory parameters (P<.05), and LASSO regression analysis screened out 10 risk factors for further study. The six independent risk factors revealed by multivariate Cox regression were age (OR 1.035, 95% CI 1.017-1.054; P<.001), CK level (OR 1.002, 95% CI 1.0003-1.0039; P=.02), CD4 count (OR 0.995, 95% CI 0.992-0.998; P=.002), CD8 % (OR 1.007, 95% CI 1.004-1.012, P<.001), CD8 count (OR 0.881, 95% CI 0.835-0.931; P<.001), and C3 count (OR 6.93, 95% CI 1.945-24.691; P=.003). The areas under the curve of the prediction model for 0.5-week, 1-week, 2-week and 3-week nonsevere probability were 0.721, 0.742, 0.87, and 0.832, respectively. The calibration curves showed that the model had good prediction ability within three weeks of disease onset. CONCLUSIONS: This study presents a predictive nomogram of critical patients with COVID-19 based on LASSO and Cox regression analysis. Clinical use of the nomogram may enable timely detection of potential critical patients with COVID-19 and instruct clinicians to administer early intervention to these patients to prevent the disease from worsening.

Comparison of clinical characteristics of COVID-19 between elderly patients and young patients: a study based on a 28-day follow-up.

The number of corona virus disease 2019 cases is increasing rapidly. However, the comparison of clinical characteristics between patients ≥ 70 and those < 70 has not been implemented yet. To achieve that, we collected clinical data of consecutive 222 patients in Renmin Hospital of Wuhan University diagnosed between January 13, 2020 and February 4, 2020. We divided them into an under-70 group and an over-70 group according to their ages, comparing their clinical characteristics. Meanwhile, univariate and multivariate Cox regression analyses were performed to identify the prognostic factors. Among the patients enrolled, 37 (16.67%) were 70 or older and 185 (83.33%) were younger than 70. Higher proportions of dyspnoea, expectoration, chronic cardiovascular disease, diabetes, organ complications, severe-to-critical cases, a higher death rate, a longer hospital stay and decreased immune status were observed in the over-70 group patients compared with their younger counterparts. The risk factors for death included dyspnoea, muscle ache, elevated myocardial enzymes, elevated C3 in over-70 patients and dyspnoea, pharyngalgia, chronic cardiac disease, increased C-reactive protein, IgA, decreased platelets in under-70 patients. Overall, our research compared the clinical characteristics of the two populations with different immune status and illustrated differentiated risk factors for death in them.

Novel and potent inhibitors targeting DHODH are broad-spectrum antivirals against RNA viruses including newly-emerged coronavirus SARS-CoV-2.

Emerging and re-emerging RNA viruses occasionally cause epidemics and pandemics worldwide, such as the on-going outbreak of the novel coronavirus SARS-CoV-2. Herein, we identified two potent inhibitors of human DHODH, S312 and S416, with favorable drug-likeness and pharmacokinetic profiles, which all showed broad-spectrum antiviral effects against various RNA viruses, including influenza A virus, Zika virus, Ebola virus, and particularly against SARS-CoV-2. Notably, S416 is reported to be the most potent inhibitor so far with an EC50 of 17 nmol/L and an SI value of 10,505.88 in infected cells. Our results are the first to validate that DHODH is an attractive host target through high antiviral efficacy in vivo and low virus replication in DHODH knock-out cells. This work demonstrates that both S312/S416 and old drugs (Leflunomide/Teriflunomide) with dual actions of antiviral and immuno-regulation may have clinical potentials to cure SARS-CoV-2 or other RNA viruses circulating worldwide, no matter such viruses are mutated or not.

A Mouse Model of SARS-CoV-2 Infection and Pathogenesis.

Since December 2019, a novel coronavirus SARS-CoV-2 has emerged and rapidly spread throughout the world, resulting in a global public health emergency. The lack of vaccine and antivirals has brought an urgent need for an animal model. Human angiotensin-converting enzyme II (ACE2) has been identified as a functional receptor for SARS-CoV-2. In this study, we generated a mouse model expressing human ACE2 (hACE2) by using CRISPR/Cas9 knockin technology. In comparison with wild-type C57BL/6 mice, both young and aged hACE2 mice sustained high viral loads in lung, trachea, and brain upon intranasal infection. Although fatalities were not observed, interstitial pneumonia and elevated cytokines were seen in SARS-CoV-2 infected-aged hACE2 mice. Interestingly, intragastric inoculation of SARS-CoV-2 was seen to cause productive infection and lead to pulmonary pathological changes in hACE2 mice. Overall, this animal model described here provides a useful tool for studying SARS-CoV-2 transmission and pathogenesis and evaluating COVID-19 vaccines and therapeutics.

The clinical course and its correlated immune status in COVID-19 pneumonia.

OBJECTIVES: To explore the clinical course and its dynamic features of immune status in COVID-19 patients and find predictors correlated with severity and prognosis of COVID-19. METHODS: The electronic medical records of 204 patients with COVID-19 pneumonia confirmed by nucleic acid testing were retrospectively collected and analyzed. RESULTS: All patients were divided into severe (69) and non-severe group (135). Lymphocyte subsets count, including CD3+ T cell, CD4+ T cell, CD8+ T cell, B cell (CD19+) and NK cell (CD16+ 56+), were significantly lower in severe group (P＜0.001). The dynamic levels of T lymphocyte in severe group were significantly lower from disease onset, but in the improved subgroup the value of T lymphocyte began to increase after about 15-day treatment and finally returned to the normal level. The cut-off value of the counts of CD3+ (576), CD4+ (391) and CD8+ (214) T cell were calculated and indicated significantly high sensitivity and specificity for severity of COVID-19. CONCLUSION: Our results shown that the decrease of CD3+, CD4+ and CD8+ T lymphocyte correlated with the course of patients with COVID-19 pneumonia, especially in severe cases. The level of T lymphocyte could be used as an indicator for prediction of severity and prognosis of patients with COVID-19 pneumonia. The application of glucocorticoid should be cautious in severe cases.

CT imaging and clinical course of asymptomatic cases with COVID-19 pneumonia at admission in Wuhan, China.

PURPOSE: Aimed to characterize the CT imaging and clinical course of asymptomatic cases with COVID-19 pneumonia. METHODS: Asymptomatic cases with COVID-19 pneumonia confirmed by SARS-COV-2 nucleic acid testing in Renmin Hospital of Wuhan University were retrospectively enrolled. The characteristics of CT imaging and clinical feature were collected and analyzed. RESULTS: 58 asymptomatic cases with COVID-19 pneumonia admitted to our hospital between Jan 1, 2020 and Feb 23, 2020 were enrolled. All patients had history of exposure to SARS-CoV-2. On admission, patients had no symptoms and laboratory findings were normal. The predominant feature of CT findings in this cohort was ground glass opacity (GGO) (55, 94.8%) with peripheral (44, 75.9%) distribution, unilateral location (34, 58.6%) and mostly involving one or two lobes (38, 65.5%), often accompanied by characteristic signs. After short-term follow-up, 16 patients (27.6%) presented symptoms with lower lymphocyte count and higher CRP, mainly including fever, cough and fatigue. The evolution of lesions on CT imaging were observed in 10 patients (17.2%). The average days of hospitalization was19.80±10.82 days, and was significantly longer in progression patients (28.60±7.55 day). CONCLUSION: CT imaging of asymptomatic cases with COVID-19 pneumonia has definite characteristics. Since asymptomatic infections as "covert transmitter", and some patients can progress rapidly in the short term. It is essential to pay attention to the surveillance of asymptomatic patients with COVID-19. CT scan has great value in screening and detecting patients with COVID-19 pneumonia, especially in the highly suspicious, asymptomatic cases with negative nucleic acid testing.