key: cord-0794348-w9kj10wo authors: Xu, Jing; Gao, Liangqin; Liang, Huiqing; Chen, Shao-dong title: In Silico Screening of Potential Anti-COVID-19 Bioactive Natural Constituents from Food Sources by Molecular Docking date: 2020-11-07 journal: Nutrition DOI: 10.1016/j.nut.2020.111049 sha: 97cc309d3bf6b082a0242c33a223eddf56b95a72 doc_id: 794348 cord_uid: w9kj10wo OBJECTIVE: Through molecular docking of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) 3CL hydrolytic enzyme (3CL(pro)) and angiotensin-converting enzyme Ⅱ (ACE2), to seek potential natural compounds that can resist Corona Virus Disease 2019 (COVID-19) using computer virtual screening technology. METHODS: Molecular docking was achieved by using the Autodock Vina software in this paper. The natural phytocompounds acting on 3CL(pro) and ACE2 were then selected from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP). Afterward, the mechanism of action of phytocompounds was speculated. RESULTS: A total of six potential natural anti- COVID-19 phytocompounds were selected, which were evaluated by the “Absorption, Distribution, Metabolism and Excretion (ADME)” and "Lipinski” rules. The content of the six phytocompounds in various fruits and vegetables was determined by the literature mining. Red wine and Chinese hawthorn and blackberry were recommended as supplements because they contained antiviral phytocompounds. CONCLUSION: Red wine and Chinese hawthorn and blackberry are theoretically proven to resist COVID-19 and are thus recommended as supplements to prevent the infection of COVID-19 during its outbreak period. The coronavirus (CoV) disease is an infectious disease caused by a newly discovered coronavirus, characterized by rapid and extensive spread, strong infectivity and general susceptibility of the population. Currently, there is no specific drug for it. The processing,and that is primarily achieved by 3C-like proteinase (3CLpro). [2] .On January 26th, professors Rao ZH and Yang HT's research team from Shanghai University of Science and Technology has obtained a high-resolution crystal structure of 2019-nCoV coronavirus 3CL hydrolase (M pro ), which is considered to be an effective target of the COVID-19 virus. [3] These studies have brought hope for the seeking of effective drugs to prevent and control the COVID-19, and may help us to develop a more effective way to fight against the COVID-19. Scientists have been focusing on searching antiviral phytocompounds with low toxicity and high curative effect from natural plants in recent years. Natural active substances have the characteristics of novel structure, high activity, and few side effects. Natural compounds from plant origin currently used in medicine exhibit chemical diversity, demonstrating their importance in modern drug discovery efforts. There is a growing trend to explore plants for pharmacologically active compounds and nutraceutical supplements. Molecular docking is a method of drug design based on the characteristics of the receptor and the way the receptor interacts with the drug molecule. As an emerging research method combining the physical and chemical principles with scientific calculation algorithms, molecular docking provides a feasible strategy for exploring the basis and mechanism of the phytocompounds [4] . This study took SARS-CoV-2 3CL pro and ACE2 as receptors, and molecular docking of the two was performed to select potential antiviral active ingredients for the development of effective and quick-acting chemical components that can resist the COVID-19. Pharmacokinetic analysis of biological or pharmaceutically active compounds was conducted to select drug candidates [5] . ADME screening criteria for ligands in this to Lipinski's rule of five will have better pharmacokinetic properties and higher bioavailability in the metabolic process in vivo, and are therefore more likely to be made into oral drugs. In this study, the phytocompounds were further chosen from the small molecule database, which was an efficient way to find compounds with good pharmacokinetic properties and high bioavailability. Ligands of this particular study were analyzed using http://www.scfbio-iitd.res.in/software/drugdesign/lipinski.jsp based on Lipinski's rule of five. According to the optimal binding energy (affinity) of phytocompounds with 3CL pro and ACE2, potential anti-COVID-19 phytocompounds were identified through ADME analysis and Lipinski screening. Based on the goal of this research to find common and easily available food supplements that can help prevent COVID-19, literature mining was conducted using PubMed, Web of Science, and EBSCO to seek the food sources of these potential phytocompounds. The principle of literature mining is that as many compounds as possible are enriched in the same food source in the hope that it can prevent viruses to a greater extent, and the food is widely distributed and easily available. In this study, SARS and viral pneumonia were used as references to search for potential targets of anti-COVID-19 phytocompounds. The potential targets of phytocompounds were predicted by the Swiss Target Prediction server. STRING database was employed to the analysis of the relationship between drugs and targets. Then, the visual analysis was carried out using Cytoscape software. Subsequently, the potential targets of the selected active components were submitted to the bioinformatics database DAVID 6.8 for functional annotation of GO genes and enrichment analysis of KEGG and REACTOME pathways, in order to further investigate the functions of these targets and their role in the signaling pathways, thereby exploring and predicting the potential molecular mechanism of phytocompounds. It is generally believed that the lower the stabilization energy of ligand binding to the receptor, the greater the possibility of action. In order to minimize the probability of false-positive results, the optimal binding energy of phytocompounds was compared with that of the currently recommended clinical chemical drugs in this study, and the binding energy in screening criteria was changed to ≤−5.0kcal/mol (-20 kJ/mol). The partial results are shown in Table 1 The molecular docking modes of six potential anti-COVID-19 phytocompounds with SARS-CoV-2 3CL pro and ACE2 are shown in Figure 1 . According to the ADME screening criteria, puerarin was eliminated due to its relatively low oral bioavailability. The remaining five plant compounds participated in further literature mining to enrich as many candidate compounds as possible in the same food source. Quercetin is a bioflavonoid widely present in red wine, grapefruit, onions, apples and black tea, having antioxidant and anti-inflammatory activity. A lesser amount of it exists in leafy green vegetables and beans [6] . Red wine alcohol is shown to be able to promote quercetin absorption and direct its metabolism towards isorhamnetin and tamarixetin [7] . Several studies have determined luteolin and isorhamnetin in red wine. In addition, [8] Quercetin and isorhamnetin are the most abundant flavonoids in plant-based foods [9] . Maja Mikulic-Petkovsek [10] detected two antiviral phytocompounds (quercetin and isorhamnetin) in 28 wild and cultivated berries (including Chinese hawthorn and blackberry) from around the world. Quercetin and isorhamnetin were found to be always present in all red-skinned species in this study. Luteolin has been identified in both Chinese hawthorn [11] and blackberry [12] in recent studies. Red wine, Chinese hawthorn and blackberry are enriched with three of the five anti-COVID-19 phytocompounds, and can be found in almost every corner of the world, so they are recommended as anti-COVID-19 food supplements. The potential targets of three bioactive natural constituents (quercetin, luteolin and isorhamnetin) and the disease targets were input into the R platform for the identification of the intersection between the two kinds of targets. The Venn diagram shows the overlap of the two kinds of targets. (Figure 3 ) The enrichment analysis of KEGG and REACTOME pathways was carried out to elucidate the critical pathways of the potential targets for quercetin and isorhamnetin in resisting the COVID-19. It was observed that 130 pathways were significantly enriched in KEGG and 172 in REACTOME. After removing the duplicates, the results were compared with the super pathways of SARS and viral pneumonia. 12 overlapping potential pathways were selected ultimately. (Fig.5 ) Fig. 5 The overlapping potential pathways selected In this study, six phytocompounds from natural plants with low binding energy to receptors were firstly selected through molecular docking. Red wine and Chinese hawthorn and blackberry are recommended as anti-COVID-19 supplements because they contain two or three active phytocompounds (i.e., quercetin, luteolin and isorhamnetin), as indicated by the extensive literature search. Reports have suggested that flavonoids, as a large class of natural compounds, might be useful for the prevention of a number of diseases, partly due to their anti-inflammatory properties [13] . Quercetin, luteolin and isorhamnetin have been shown to have antioxidant and anti-inflammatory effects in previous studies [6, 14, 15] . The results of a recent study showed that quercetin supplementation reduced all pathological changes in mice with rhinovirus-induced chronic obstructive pulmonary disease (COPD) and might prevent pulmonary disease progression in COPD. [16] Another experiment confirmed that quercetin could enhance ligand-induced senescent idiopathic pulmonary fibrosis fibroblast apoptosis and reduce lung fibrosis in vivo. [17] Luteolin has been proved to improve experimental pulmonary fibrosis in vivo and in vitro [18] and attenuate acute lung injury in experimental mouse models [19, 20] . Isorhamnetin protects mice from acute lung injury by suppressing inflammation, [21] and prevents bleomycin-induced pulmonary fibrosis by inhibiting endoplasmic reticulum stress and epithelial-mesenchymal transformation. [22] To further check the anti-COVID-19 mechanism of three active phytocompounds, the network pharmacology tool was used to analyze the targets of the phytocompounds, the cell signal transduction pathways that might be involved in the regulation, and the potential pharmacological mechanism. SARS and viral pneumonia were used as reference diseases. Almost half of the targets of each phytocompound intersected with the targets of SARS and viral pneumonia. The results of network pharmacologic analysis indicated that the targets of the three plant compounds might resist the COVID-19 through 12 overlapping pathways, three of which (the small-cell lung cancer pathway, non-small cell lung cancer, and tuberculosis) acted directly on the lungs. Because of the mechanisms of immune resistance and tissue resilience, the Innate Immune System pathway plays an important role in the worsened pneumonia in a subset of patients [23] . An earlier study confirmed that flavonoids inhibited cell proliferation and induced apoptosis and autophagy through downregulation of PI3Kγ-mediated PI3K-Akt signaling pathway [24] . It has been found in many studies that drugs can suppress the inflammatory response through the PI3K-Akt signaling pathway. [25] [26] [27] [28] TNF drives the release of itself and other proinflammatory cytokines (e.g., IL-1β and IL-6) [29, 30] and participates in the systemic inflammatory response. It is one of the cytokines that contribute to the acute phase response. A study revealed that spatial heterogeneity of the T cell receptor repertoire reflected the mutational landscape in lung cancer [31] , and T cell-targeted immunotherapy has been increasingly applied to the treatment of non-small cell lung cancer [32] . The enrichment of these pathways from network pharmacology confirmed the hypothesis that quercetin, luteolin and isorhamnetin were natural and effective anti-COVID-19 supplementaries. Given the predictability of the virtual screening results, further in vitro and in vivo experiments are needed to verify the results of this study if possible, so as to provide the experimental basis for the development of natural anti-COVID-19 supplementaries. Unfortunately, many experiments couldn't be carried out due to the current epidemic, so this research was more based on previous studies and theoretical analysis. In addition, due to the human metabolism, the concentration of active ingredients in food is difficult to reach the level of clinical drugs, and the preventive effect of food may be relatively weak. Red wine, Chinese hawthorn and blackberry are rich dietary sources of polyphenols with reported health benefits. They are recommended as preventive supplements because they contain three anti-COVID-19 phytocompounds (i.e., quercetin, luteolin and isorhamnetin). The analysis of the anti-COVID-19 mechanism of these three compounds by Internet pharmacology tools identifies several key pathways, which theoretically confirms the hypothesis that berries are a natural anti-COVID-19 supplement. The authors declare that they have no competing interests. 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Material preparation, data collection performed by JX and SDC, and analysis were performed by JX and LQG.The first draft of the manuscript was written by JX and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Supported by the Fundamental Research Funds for the Central Universities