Objective: ´Three formulas and three medicines,’ namely, Jinhua Qinggan Granule, Lianhua Qingwen Capsule, Xuebijing Injection, Qingfei Paidu Decoction, HuaShi BaiDu Formula, and XuanFei BaiDu Granule, were proven to be effective for coronavirus disease 2019 (COVID-19) treatment. The present study aimed to identify the active chemical constituents of this traditional Chinese medicine (TCM) and investigate their mechanisms through interleukin-6 (IL-6) integrating network pharmacological approaches.
Methods: We collected the compounds from all herbal ingredients of the previously mentioned TCM, but those that could down-regulate IL-6 were screened through the network pharmacology approach. Then, we modeled molecular docking to evaluate the binding affinity between compounds and IL-6. Furthermore, we analyzed the biological processes and pathways of compounds. Finally, we screened out the core genes of compounds through the construction of the protein–protein interaction network and the excavation of gene clusters of compounds.
Results: The network pharmacology research showed that TCM could decrease IL-6 using several compounds, such as quercetin, ursolic acid, luteolin, and rutin. Molecular docking results showed that the molecular binding affinity with IL-6 of all compounds except γ-aminobutyric acid was < −5.0 kJ/mol, indicating the potential of numerous active compounds in TCM to directly interact with IL-6, leading to an anti-inflammation effect. Finally, Cytoscape 3.7.2 was used to topologize the biological processes and pathways of compounds, revealing potential mechanisms for COVID-19 treatment.
Conclusion: These results indicated the positive effect of TCM on the prevention and rehabilitation of COVID-19 in at-risk people. Quercetin, ursolic acid, luteolin, and rutin could inhibit COVID-19 by down-regulating IL-6.
In December 2019, in Wuhan, Hubei Province, China, the Chinese Center for Disease Control and Prevention identified a highly contagious novel coronavirus (SARS-CoV-2) [
One of central challenges for COVID-19 treatment is the myriad of proinflammatory cytokines released during the disease progression, known as a cytokine release syndrome (CRS) [
Patients with severe COVID-19-associated pneumonia may exhibit systemic hyperinflammation, known as macrophage activation syndrome or cytokine storm [
Traditional Chinese medicine (TCM) has been proven effective for COVID-19 treatment [
Our research aimed to systematically investigate the active components of ‘three formulas and three medicines’ for COVID-19 treatment and the mechanism based on IL-6 integrating network pharmacological methods.
Supplementary Table S1 outlines Jinhua Qinggan Granules, Lianhua Qingwen Capsules, Xuebijing Injection, Qingfei Paidu Decoction, XuanFeiBaiDu Granule, and HuaShi BaiDu Formula, which were obtained from traditional medicine guidelines and Chinese Clinical Trial Registry up to April 30, 2020. The information on the active compounds of these ingredients in TCM was downloaded from the Traditional Chinese Medicine Systems Pharmacology (TCMSP,
Pharmacokinetic parameters (chemical structure; oral bioavailability (OB); drug-likeness (DL); blood–brain barrier (BBB) permeability; half-life of compounds) were obtained from the TCMSP database and confirmed by the DrugBank (
CRS is the main cause of morbidity in SARS-CoV and MERS-CoV patients [
The information on the active compounds that interact with IL-6 was downloaded from the TCMSP database, and then we identified and confirmed those that could down-regulate IL-6 [
In recent years, based on bioinformatics and systems biology, network pharmacology has been applied in many fields of life sciences, such as in the identification of new drug targets, compound discovery, and evaluation of preclinical efficacy. The thinking method of integrated regulation of multiple targets based on molecular docking, construction of drug–target network, and analysis of network characteristics is used gradually in predicting the main active components and potential targets of TCM and elaborating the mechanism of TCM. To further characterize the molecular mechanism and topological structure of the medicine/formula, compounds, and IL-6, interaction networks were built and visualized using Cytoscape 3.7.2 (
ChemOffice software was used for the construction of the 3D structures of the chemical constituents of TCM. Then, MMFF94 force field was used to minimize the energies of chemical constituents. The RCSB Protein Data Bank (PDB) was used to obtain the 3D structure of IL-6 in PDB format [
To understand the biological processes and pathways of TCM in COVID-19 treatment, Cytoscape 3.7.2 was used to analyze the biological processes and pathways of their compounds. It integrates several authoritative databases, such as GO, KEGG, and DrugBank, hence providing researchers with comprehensive and detailed information on genes. The genes of compounds were obtained from the TCMSP database and confirmed by the DrugBank database. All the gene names were standardized through the UNiProtKB (
Gene clusters are groups of genes with the same or similar functions, which are closely related in some biological processes and pathways. We then intended to identify the hub genes in compound-related genes.
Active compounds that interact with IL-6 were retrieved from the TCMSP database, and then we identified and confirmed those that could down-regulate IL-6.
The Herbs–Compounds–IL-6 network was constructed using Cytoscape 3.7.2. The complex relationship between each other could be observed in this network. The blue hexagon represents herb, and the red diamond represents compound. The green V represents IL-6.
Compound | OB(%) | DL | BBB | HL | RBN |
---|---|---|---|---|---|
Rutin | 3.20 | 0.68 | -2.75 | - | 6 |
Matrine | 63.77 | 0.25 | 1.52 | 6.69 | 0 |
Melanin | 26.40 | 0.67 | -0.63 | - | 0 |
Luteolin | 36.16 | 0.25 | -0.84 | 15.94 | 1 |
Caffeine | 89.46 | 0.08 | -0.01 | 13.64 | 0 |
Aucubin | 4.17 | 0.33 | -2.90 | - | 4 |
Piperine | 42.52 | 0.23 | 0.62 | 10.25 | 3 |
Wogonin | 30.68 | 0.23 | 0.04 | 17.75 | 2 |
Daidzein | 19.44 | 0.19 | -0.22 | - | 1 |
Myricetin | 13.75 | 0.31 | -1.01 | - | 1 |
Quercetin | 46.43 | 0.28 | -0.77 | 14.40 | 1 |
Ursolic acid | 16.77 | 0.75 | 0.07 | - | 1 |
Resveratrol | 19.07 | 0.11 | -0.01 | - | 2 |
Paeoniflorin | 53.87 | 0.79 | -1.86 | 13.88 | 7 |
Sinomenine | 30.98 | 0.46 | 0.43 | 1.79 | 2 |
Atractylenolide I | 37.37 | 0.15 | 1.29 | 7.10 | 0 |
γ-aminobutyric acid | 24.09 | 0.01 | -0.57 | - | 3 |
Abbreviations: BBB, blood–brain barrier; DL, drug-likeness; HL, half-life; OB, oral bioavailability; RBN, rotatable bond number.
We then investigated whether TCM could suppress IL-6.
Consistence with
Jinhua Qinggan granules | Lianhua Qingwen capsules | Xuebijing Injection | Qingfei Paidu Decoction | XuanFeiBaiDu Granule | HuaShiBaiDu Formula | |
---|---|---|---|---|---|---|
Rutin | √ | √ | √ | √ | √ | |
Matrine | ||||||
Melanin | ||||||
Luteolin | √ | √ | √ | √ | √ | |
Caffeine | ||||||
Aucubin | √ | |||||
Piperine | ||||||
Wogonin | √ | √ | √ | |||
Daidzein | ||||||
Myricetin | √ | √ | √ | |||
Quercetin | √ | √ | √ | √ | √ | √ |
Ursolic acid | √ | √ | √ | √ | √ | √ |
Resveratrol | √ | |||||
Paeoniflorin | √ | √ | ||||
Sinomenine | ||||||
Atractylenolide I | √ | |||||
γ-aminobutyric acid | √ | √ |
Quercetin, Ursolic acid, Luteolin, Rutin, and so on could be found in medicines and formulas. Quercetin, Ursolic acid, Luteolin, and Rutin were the most frequently used compounds in these medicines and formulas.
To obtain the best combination scheme of Chinese herbs, we made an alluvial diagram of Chinese herbs pertaining to the above-mentioned compounds.
The result showed that Forsythiae Fructus (rutin, luteolin, aucubin, wogonin, myricetin, quercetin, and ursolic acid), Lonicerae Japonicae Flos (rutin, luteolin, quercetin, and ursolic acid), Carthami Flos (rutin, luteolin, myricetin, and quercetin), and Verbenae herb (luteolin, aucubin, quercetin, and ursolic acid) had the most abundant compounds.
In summary, first, these results suggested that TCM could have a therapeutic effect by reducing IL-6 (
The results indicated that they could exert therapeutic effects by suppressing IL-6 production. The hexagon represents herb, and the blue diamond represents compound. The red triangle represents IL-6.
To know whether the chemical constituents of TCM will interact directly with IL-6, molecular docking was modeled, evaluating the binding affinity between them.
Schematic diagrams demonstrating the IL-6-binding sites and the proximate affinity of candidate compounds in TCM.
Compound | Chemical formula | Molecular weight | Binding affinity (kJ/mol) |
---|---|---|---|
Rutin | C27H30O16 | 610.517 | -5.9 |
Matrine | C15H24N2O | 248.364 | -7.0 |
Melanin | C18H10N2O4 | 318.283 | -7.1 |
Luteolin | C15H10O6 | 286.236 | -8.1 |
Caffeine | C8H10N4O2 | 194.191 | -5.5 |
Aucubin | C15H22O9 | 346.33 | -5.7 |
Piperine | C17H19NO3 | 285.338 | -5.7 |
Wogonin | C16H12O6 | 300.263 | -7.3 |
Daidzein | C15H10O4 | 254.237 | -5.7 |
Myricetin | C15H10O8 | 318.235 | -8.2 |
Quercetin | C15H10O7 | 302.236 | -8.2 |
Ursolic acid | C30H48O3 | 456.7 | -6.4 |
Resveratrol | C14H12O3 | 228.243 | -6.5 |
Paeoniflorin | C23H28O11 | 480.462 | -6.1 |
Sinomenine | C19H23NO4 | 329.39 | -6.7 |
Atractylenolide I | C15H18O2 | 230.302 | -6.4 |
γ-Aminobutyric acid | C4H9NO2 | 103.12 | -3.1 |
All compounds except γ-aminobutyric acid molecular binding affinity with IL-6 were less than −5.0 kJ/mol, indicating that several bioactive chemical compounds in TCM have the potential to interact directly with IL-6.
We downloaded the information on compound-related genes from the TCMSP database (
ClueGO plug-in was used to analyze the interaction networks of enriched biological processes and pathways. The multiple color dots show that it revolved in multiple biological processes and pathways.
Compound | Target genes |
---|---|
Matrine | TNF; IL-6 |
Paeoniflorin | TNF; IL-6 |
Atractylenolide I | IL1B; CD40LG; IL6; GABRA1; PGF; VEGFA |
Resveratrol | CA2; HSP90AA1; PTGS1; PTGS2; MAOB; NCOA2 |
Melanin | PTGS1; PTGS2; MAPK3; MAPK1; VEGFA; TNF; IL-6 |
Aucubin | SERPIND1; CA2; PTGDR2; DPP4; CD40LG; BCL2; IL6; FAM213B |
Caffeine | MAPK1; SLC6A2; TP53; GRIA2; ADRB2; PDE4B; ADORA2A; CYP1A2; KCNJ11 |
Rutin | CAT; IL1B; SOD1; HMGCR; CD40LG; ALOX5; INS; ITGB2; GSTP1; IL6; TBXA2R |
Myricetin | MMP2; PROC; IL1B; AKR1B1; PPARG; PCK1; HSP90AA1; DPP4; TOP1; PTGS1 |
Ursolic acid | MMP2; CDK4; IL1B; FGF2; CDK6; TP53; MMP10; MMP3; FASN; MMP1; SELE |
Piperine | LTA4H; IL12B; CHRM3; IL1B; SLC6A3; ADRB2; CHRM1; ADRA1B; MAOA PDE3A; PTGS2; SOAT1; CD40LG; MAOB; IL6; ADRA2C; SCN5A; ABCB; ENSG00000196689 |
Sinomenine | SLC6A2; IL2; IFNG; OPRD1; CHRM3; ADRB2; CHRM1; ADRA1B; HSP90AA1 |
Luteolin | MAPK1; HMOX1; MMP2; IL2; IFNG; CDK4; RB1; TP53; INSR; PRSS1; |
\Wogonin | CCL2; MAPK14; KDR; CDK2; TP53; PPARG; ADRB2; PRSS1; MMP1; GSK3B |
Daidzein | MAPK14; CAT; AHR; ATP5B; CDK2; TP53; PPARG; VCAM1; ADRB2; PRSS1 |
Quercetin | SERPIND1; MAPK1; HMOX1; MMP2; PLAT; PON1; PON2; MPO; CCL2; COL1A1 |
These genes were downloaded from TCMSP database and confirmed by Drugbank database. All the genes names were standardized through UNiProtKB database with the ‘Homo sapiens’.
Construction of PPI network and excavation of gene clusters of compounds to sieve the core genes of compounds, we uploaded the genes in
We divided each PPI network into Model A to Model D based on their cluster score; genes in Model A cluster had the highest score for each compound.
Compound | Model A | Model B | Model C | Model D |
---|---|---|---|---|
Rutin | 4.500 | 3.000 | / | / |
Melanin | 6.333 | / | / | / |
Luteolin | 9.800 | 4.000 | / | / |
Caffeine | 7.000 | 3.000 | / | / |
Piperine | 5.000 | / | / | / |
Wogonin | 11.538 | / | / | / |
Daidzein | 11.833 | 3.000 | / | / |
Myricetin | 7.111 | / | / | / |
Quercetin | 18.700 | 4.000 | 3.500 | 3.000 |
Ursolic acid | 13.571 | / | / | / |
Sinomenine | 5.000 | 3.333 | / | / |
In December 2019, in Wuhan, Hubei Province, China, some hospitals successively found a number of pneumonia cases of an unknown cause with a history of exposure to the South China seafood market, which has now been confirmed as an acute respiratory infectious disease caused by a novel coronavirus (COVID-19).
Although several western medicines exerted protective effects on vulnerable people [
However, the potential mechanism of these medicines remains poorly understood. Emerging evidence indicates that high levels of IL-6 are observed in COVID-19 patients [
As a naturally occurring flavonoid, quercetin is widely distributed in every part of the plant, such as the roots, stems, leaves, flowers, and fruits. Quercetin has a wide range of pharmacological effects on many diseases. A growing body of evidence indicates that the alleviation of diabetic encephalopathy and protection of human oral keratinocytes could be seen in quercetin through antioxidant, anti-inflammatory, and antiapoptotic effects [
Ursolic acid (UA) was the second common compound in the present study. UA is a pentacyclic triterpenoid compound, which is widespread in plants [
Like quercetin, luteolin is also a common antioxidant [
Rutin is one of the classic flavonoids. Several studies have investigated its anti-tumor and anti-inflammatory effects [
In summary, quercetin, UA, luteolin, and rutin were polyphenols extracted from plants that have a wide range of biological effects, including anti-carcinogenic, anti-inflammatory, and anti-viral, which reduce lipid peroxidation, platelet aggregation, and capillary permeability. The compounds from the ‘three formulas and three medicines’ could hopefully treat COVID-19. The limitation of this manuscript is the lack of clinical trials on these compounds for COVID-19 treatment.
Our study indicated that several compounds such as quercetin, UA, luteolin, and rutin could decrease IL-6 expression, showing an anti-CRS effect in COVID-19 patients.
Click here for additional data file.
The data used to support the findings of the present study are included within the article. Any further data can be made available from the corresponding author upon request.
The authors declare that there are no competing interests associated with the manuscript.
This work was supported by the National Natural Science Foundation of China (NSFC) [grant numbers 81270405, 91539118, and 81611130092]; Program of Shanghai Academic Research Leader [grant number 17XD1405000 (to C.L.)]; and NSFC [grant number 81503371 (to F.W.) and 81801095 (to Y.-C.C.)].
Wen-hao Niu and Feng Wu designed the study. Wen-yue Cao analyzed the data. Wen-yue Cao, Yu-Chieh Chao, and Zong-gui Wu provided support to this research. Wen-hao Niu and Feng Wu wrote this study. Chun Liang and Fei Peng revised this manuscript.
blood–brain barrier
coronavirus disease 2019
drug-likeness
half-life
interleukin-6
oral bioavailability
protein–protein interaction
rotatable bond number
traditional Chinese medicine