Source: Frontiers in Life Sciences
Glycyrrhizin is an important monomer active ingredient in licorice, which has a variety of pharmacological effects such as antioxidant and anti-HIV. It has an inhibitory effect on the ulcer formed by pyloric ligation in rats, and can produce morphological changes on rat ascites liver cancer and mouse Ascites carcinoma cells.
On May 2, researchers at Peking University and the Academy of Military Medical Sciences screened a compound from 3682 compounds, liquiritin (CAS number: 551-15-5), which can effectively inhibit SARS-CoV-2 infection, through a new strategy, and the results were published in the uncommitted platform bioRxiv.
Compound screening
Using the previously reported transcriptome data of AT2 cells expressing the new coronavirus receptor ACE2 as a reference, the researchers selected genes related to the viral processing process, combined with gene collection and enrichment analysis (GSEA) means and artificial intelligence pharmacodynamic prediction platform Infinity Phenotype (related articles are being published) to simulate the drug candidates, and found that glycyrrhizin had the highest enrichment score (ES score) in the simulation analysis of antiviral-related gene differential expression.
Licoriceside antiviral test
To verify the screening results of the above predictions, the researchers tested the effects of glycyrrhizin on SARS-CoV-2 virus yield and infection efficiency in Vero E6 cells. First, cytopathic effectivity (CPE) inhibition experiments showed that 10 μM glycyrrhizin showed a significant CPE inhibitory effect; secondly, the micromolal concentration (μM) glycyrrhizin could effectively inhibit viral infection, and half of the maximum effect concentration EC50 was only 2.39 μM.
Exploration of the mechanism of glycyrrhizin antiviral
To further shed light on the mechanisms by which glycyrrhizins fight viruses, the researchers borrowed transcriptome data from MCF7 cells after glycyrrhizin treatment, published in 2017.
GO analysis of the data (functional enrichment analysis of differentially expressed genes) showed that glycyrrhizin can upregulate gene expression in the following functional aspects—genes in the interferon I signaling pathway, negatively regulated viral genome replication genes, defense virus response genes, and viral response genes (Figure b below);
At the same time, inflammation-related gene expression was also down-regulated, suggesting that glycyrrhizin reduces the inflammatory response (Figure C below). Among the upregulated genes, multiple genes associated with the type I interferon signaling pathway were significantly upregulated (the most different, in the first group, figure b below), including interferon downstream genes.
These genes form an interacting network (Figure d below) that suggests that glycyrrhizin exerts its antiviral activity by mimicking type I interferons (or their pathways).
Previously, Peking University Hongbo Liu reported that brass derivatives can effectively inhibit 3CLpro protease activity (3CLPro is a key enzyme for SARS-CoV-2 replication, and viral polypsins can be cleaved by 3CLPro into multiple active proteins such as viral replication protein RdRp).
Glycyrrhizin, also a flavonoid derivative, has a parent nucleus structure similar to the compound reported by Hongbo Liu et al., and whether it also has an effect on 3CLpro.
Although the molecular docking model showed that glycyrrhizin can bind to 3CLpro, the authors' in vitro experiments did not find that glycyrrhizin inhibited the enzymatic activity of 3CLpro (the relevant data were not presented by the authors).
Glycyrrhizin safety assessment
Firstly, the toxicity of glycyrrhizin to Vero E6 cells was assessed by MTS experiments (detection of cell proliferation), and the results showed that the concentration of glycyrrhizin at the time of death of half of the cells was CC50=29.46 μM, which was far lower than its viral semi-inhibitory concentration of 2.39 μM; the maximum overall inhibition rate of cell growth was 70%.
Secondly, after the one-time intravenous injection of 150 mg/Kg glycyrrhiside in mice, no death was found in mice during the 14-day monitoring period, and no signs of toxicity (normal body weight, food intake, and drinking water) were observed; after giving a higher dose of 300 mg/Kg of glycyrrhizin for seven consecutive days, the indicators of mice were similar to the previous dose.
At the same time, the authors measured multiple organ indexes and serum biochemical indices of the above experimental mice.
Except for the liver, the organ index (the ratio of organ weight to body weight) was not significantly different from that of the control mice; the blood glucose, cholesterol, triglycerides, and hdL lipoproteins of the mice also remained normal; and there were no abnormalities in blood urea, uric acid, creatine kinase, and α-oxybutyrate dehydrogenase, and these data showed that the kidney function and heart function of the mice were also not impaired.
Other related metrics are also within the reference values. The above results show that licoriceside has no significant toxicity to mice. (More metric data is presented in tabular form in the paper and will not be listed here)
Pharmacokinetic study of licorice glycyrrhiside
By consulting some literature studying the pharmacokinetics of glycyrrhizin mixtures, the authors found that the corresponding glycyrrhizin content in the mixture ranged from 225 μg/kg to 44000 μg/kg, the maximum concentration (Cmax) in animal serum ranged from 0.064 to 2.3 μM, and the maximum residence time (Tmax) ranged from 0.25 to 0.6 hours;
The half-life of the drug (T1/2) ranges from 1.9 to 11.63 hours. In this author, the authors propose that in view of the low biological activity and rapid degradation of licorice glycyrrhizin, it is best to choose intravenous or inhaled administration.
Whether licorice glycyrrhiside can safely and effectively resist the new crown virus infection requires more scientific experiments, such as evaluating its efficacy in patients with new crown pneumonia. Admittedly, this article provides us with new ideas for (antiviral) drug screening, providing new possibilities for the treatment of new crown pneumonia, and the authors also point out that some related follow-up experiments are underway. Don't have a double yellow farce!
bibliography
1 Zhu, J. et al. An artificial intelligence system reveals liquiritin inhibits SARS-CoV-2 by mimicking type I interferon. bioRxiv, 2020.2005.2002.074021, doi:10.1101/2020.05.02.074021 (2020).
2 Zhao, Y. et al. Single-cell RNA expression profiling of ACE2, the receptor of SARS-CoV-2. bioRxiv, 2020.2001.2026.919985, doi:10.1101/2020.01.26.919985 (2020).