# Novel Drug Design for Treatment of COVID-19: A Systematic Review of Preclinical Studies

Authors: Sarah Mousavi; Shima Zare; Mahmoud Mirzaei; Awat Feizi
Journal: Canadian Journal of Infectious Diseases and Medical Microbiology
 (2022)

---

## Abstract

Background. Since the beginning of the novel coronavirus (SARS-CoV-2) disease outbreak, there has been an increasing interest in discovering potential therapeutic agents for this disease. In this regard, we conducted a systematic review through an overview of drug development (in silico, in vitro, and in vivo) for treating COVID-19. Methods. A systematic search was carried out in major databases including PubMed, Web of Science, Scopus, EMBASE, and Google Scholar from December 2019 to March 2021. A combination of the following terms was used: coronavirus, COVID-19, SARS-CoV-2, drug design, drug development, In silico, In vitro, and In vivo. A narrative synthesis was performed as a qualitative method for the data synthesis of each outcome measure. Results. A total of 2168 articles were identified through searching databases. Finally, 315 studies (266 in silico, 34 in vitro, and 15 in vivo) were included. In studies with in silico approach, 98 article study repurposed drug and 91 studies evaluated herbal medicine on COVID-19. Among 260 drugs repurposed by the computational method, the best results were observed with saquinavir (n = 9), ritonavir (n = 8), and lopinavir (n = 6). Main protease (n = 154) following spike glycoprotein (n = 62) and other nonstructural protein of virus (n = 45) was among the most studied targets. Doxycycline, chlorpromazine, azithromycin, heparin, bepridil, and glycyrrhizic acid showed both in silico and in vitro inhibitory effects against SARS-CoV-2. Conclusion. The preclinical studies of novel drug design for COVID-19 focused on main protease and spike glycoprotein as targets for antiviral development. From evaluated structures, saquinavir, ritonavir, eucalyptus, Tinospora cordifolia, aloe, green tea, curcumin, pyrazole, and triazole derivatives in in silico studies and doxycycline, chlorpromazine, and heparin from in vitro and human monoclonal antibodies from in vivo studies showed promised results regarding efficacy. It seems that due to the nature of COVID-19 disease, finding some drugs with multitarget antiviral actions and anti-inflammatory potential is valuable and some herbal medicines have this potential.


---

## Body

## 1. Introduction

Coronavirus disease 2019 (COVID-19), which was first identified in December 2019, and shortly after, declared a pandemic by World Health Organization (WHO) [1]. As of January 18, 2022, there have been more than 326 million confirmed cases and 5.54 million deaths globally [2]. Coronaviruses belong to the family of Coronaviridae, RNA viruses with crown-like spikes on the surface of the coronavirus particles. According to a meta-analysis of Macedo et al. [3], the mortality rate of COVID-19 was 17.1% for patients admitted to hospitals, whereas WHO estimated a fatality rate of 6.73%, which was much lower than that calculated from published studies. Among the critical cases of COVID-19, the mortality rate reaches 40% [4].Substantial efforts have been made in the treatment of patients with COVID-19. The WHO recommendations in the treatment of COVID-19 are as follows [5]: molnupiravir (conditional), baricitinib (strong), ruxolitinib and tofacitinib (conditional), sotrovimab (conditional), casirivimab and imdevimab (conditional), IL-6 receptor blockers (tocilizumab and sarilumab) (strong), remdesivir (conditional), and systemic corticosteroids (strong). The WHO recommends not to use ivermectin, lopinavir/ritonavir, hydroxychloroquine, and convalescent plasma.The pathogenesis of COVID-19 was explained by cytokine storm, reduction in ACE2 expression, and activation of complement pathway-induced microvascular injury and thrombosis [6]. The mechanisms of the recommended agents are focused on the mentioned pathogenesis to improve the clinical outcome of COVID-19, and antiviral therapies are missing. The anticoronaviral strategies include preventing the synthesis of viral RNA, inhibiting virus replication, blocking the virus binding to human cell receptors, or inhibiting the viruses’ self-assembly process [7]. The SARS-CoV-2 contains at least four structural proteins: spike (S) protein, envelope (E) protein, membrane (M) protein, and nucleocapsid (N) protein, and 16 nonstructural proteins (NSPs). Among the translated NSPs, the main protease, also called chymotrypsin-like protease (3C-like protease), and the papain-like protease are two essential proteases for proteolytic processing of the coronavirus replicase polyprotein, therefore generating functional replication complex of the virus, whereas RNA-dependent RNA polymerase is the central enzyme for RNA synthesis. These three NSPs play crucial roles in coronavirus replication, making them attractive targets for anticoronaviral drug design [8].The S protein, a surface-located trimeric glycoprotein of coronaviruses, promotes the attachment of viruses to host cells through binding to angiotensin-converting enzyme 2 (ACE2) and virus-cell membrane fusion during viral infection. Thus, the S protein has been considered as a major target for the development of vaccines and drug [9].The development of a new therapeutic agent is a complex, lengthy, and expensive process, which can take 2–4 years of preclinical development and 3–6 years of clinical development and over 500 million dollar cost. There are three critical steps to develop a new drug including discovery and development, preclinical research, and clinical development [10].Drug discovery involves screening hits, medicinal chemistry, and optimization of hits to reduce potential drug side effects. For drug discovery, two different complementary approaches can be applied: classical pharmacology, also known as phenotypic drug discovery, which is the historical basis of drug discovery, and reverse pharmacology, also known as designated target-based drug discovery. Screening methods based on phenotypic drug discovery have been used to discover new natural products mainly from the terrestrial origin [11]. These two strategies have advantages and disadvantages and promote very different screening assays. The frequent re-discovery of the same compounds, the technical difficulties associated with the isolation of compounds from extracts, and the incompatibility of natural product extracts with high-throughput screening (HTS) campaigns were the disadvantages of phenotypic drug discovery. On the other hand, natural product structures have the characteristics of high chemical diversity, biochemical specificity, and other molecular properties that make them favorable as lead structures for drug discovery, which serve to differentiate them from libraries of synthetic and combinatorial compounds [12]. Overly simplified assays, acting of drug on more than one target, the multifactorial nature of diseases, and challenges to identify a single molecular target are some limitations of target-based drug discovery. Therefore, a comprehensive screening strategy will incorporate both targeted and phenotypic assays, with one format designated as the primary screen and the other as a secondary or follow-up assay. During the spread of COVID-19 outbreak, great efforts have been made in therapeutic drug discovery against the virus. Because COVID-19 is a new, acute, severe infectious disease, the anti-SARS-CoV-2 drug development strategies are to screen existing drugs to identify potentially effective drugs, to expand indications, and to develop a vaccine [12]. The safety of conventional drugs has been mostly verified; if effective, they can be quickly applied in clinical practice (repurposing of existing drugs). The recent rises of several high transmissible strains sounded alarms for currently used vaccines and drugs. Therefore, developing broad-spectrum antiviral drugs not only to combat COVID-19 but also to provide protective arsenals against future viral outbreaks is a requirement. Scientists continue the development of broad-spectrum antiviral drugs from natural or chemical sources, which have the potential advantages of broad-spectrum therapeutic effect and insensitivity to viral evasion. Given the urgency of the SARS-CoV-2 outbreak, here we discuss the discovery and development of new therapeutics for SARS-CoV-2 infection based on the strategies from preclinical drug discovery.


## 2. Methods

We report this systematic review based on the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines [13].


### 2.1. Data Sources and Searches

Studies published in PubMed, Scopus, Web of Science, EMBASE, Google Scholar, and DrugBank were searched from December 2019 to March 2021 using the following search terms: “Coronavirus,” “Covid-19,” “SARS-CoV-2,” “Drug design,” “Drug development,” “In silico,” “In vitro,” and “In vivo” alone or in combination without language restrictions. The keywords were selected using expert opinions, mesh, and related article titles.All articles with full text or in the absence of full text with abstract are included in the screening of this study. Studies were excluded if studies were comments, editorial, letters, review, and preprints.


### 2.2. Data Extraction

Two researchers independently extracted data from included studies using a predefined data extraction form. All disagreement was discussed and solved after rechecking the source data with a third investigator. The data extracted, including the last name of the first author, type of the study (in silico, in vitro, in vivo), and name of the agent (chemical compound, drug, herb, etc.), studied the mechanism and efficacy of the agent according to study design that specified according to the following definition.Computer-aided drug design can be divided into three different categories. All are based on ligands and receptors, which are briefly described [14] as follows:(1)
Dock Receptor-Based Approach [15]. Once the three-dimensional structure of the ligand molecules and their receptor is known, the receptor-based method is a good candidate for identifying or optimizing drugs. Due to the presence of three-dimensional structures of compounds and receptors, the nature of the interaction between the ligand and the receptor and the type of structure that the ligand can have to interact with them in favorable conditions can be identified using this method. The compound is simulated on the active site of the dock (meaning anchoring) and on the interaction of the ligand with the receptor by molecular mechanics and molecular dynamics. In this method, due to the ligation of the ligand in the active position, the ligand changes in terms of conformity and changes its position in different conditions and shows interaction with the receptor in different types of situations. To determine the type of ligands that can be docked into the receptor site, the matching of the shape and the complementarity of the hydrophobic, hydrophilic, and charged parts must be considered. Various software packages such as AUTODOCK-Glide-LUDI and LigandFit are used to design the drug based on the structure of the receptor.(2)
Ligand-Based Approach [16]. This method is used where the three-dimensional structures of the receptor are unknown and instead the structure of the ligands is known, which is one of the common methods. In this method, by indirectly studying compounds that react with biomolecules, they seek to design compounds that are pharmacologically active. In ligand-based drug design methods, in the absence of biomolecule structure, by studying specific ligands, it seeks to identify the structural and physicochemical properties of the compounds so that the desired compound can be designed based on data extracted from the study of previous compounds. This method is a kind of drug design based on pharmacophore (pharmacophore refers to the part of the drug to which the effect of the drug depends on that part of the molecule), and by studying the quantitative relationship between structure and their activity, drugs can be designed by this method. It can be said that it is a method for designing the pharmacophores of drugs.(3)
Denovo Design-Based Approach [17]. This method is used when the structure of the ligand is unknown but the structure of the receptor is known. In this method, there is information about the structures of the receptor or quasi-receptors, but there is no structure of the main composition that can interact with the active site of the receptor. One of the functions of drug design based on this method is to suggest and present the main composition that is complementary to the active site. The basis of the method is that the database of existing 3D structures is used to find small molecules that can interact with the active site of the receptor in terms of size, geometry, and functional groups. Software packages such as GROW and LEGEND are used to design drugs by this method.Drug design methods in the computer include quantitative structure-activity relationship (RASQ), docking, molecular dynamic simulation, and computational modeling. In these studies, the efficacy is evaluated based on the function of the drug or compound agent and the mechanism of action. Figure1 shows the methods of computer-aided drug design (CADD).Figure 1
Computational methods in drug design (CADD: computer-aided drug design).


#### 2.2.1. Computational Methods in Drug Design

Quantitative Structure-Activity Relationship (QSAR). QSAR provides studies on the relationship between chemical structure and biological activity or other biological activities that are important in selecting or removing a compound before synthesis and testing. QSAR [18] is especially important to predict the result, especially when it is not possible to experiment with a compound. Molecular descriptors, which are the most important components of QSAR, can be obtained experimentally or through mathematical formulas from various theories such as quantum mechanics, chemical graph theory, and study theories. QSAR seeks to establish a statistically significant relationship between structure and performance. It also explains the specific effect of a drug and can ultimately predict the effect of newly synthesized chemical compounds. QSAR model is also an equation that predicts a property through molecular descriptors and their coefficients. Evaluation of the effectiveness of new compounds that have been studied using this method can be reported as a percentage of enzyme inhibition if the modeling has been done and mentioned in the article.


#### 2.2.2. Docking

In this technique, to achieve a combination with a pharmacological effect and increase the pharmacological activity of the drug, different formulations of a drug interact with the receptor, and the structure that has the best interaction with the receptor and the lowest energy level is selected for laboratory steps [19]. In this way, possible structures that have a stronger interaction with the receiver can be isolated at this stage. The same issue is considered and reported as an effectiveness measure.


#### 2.2.3. Molecular Dynamic Simulation

In this technique, which is based on the simulation of drug-receptor interaction in the body, docking problems are solved and in fact play a complementary role in this technique [20]. Due to the time-consuming work with this technique, effective structures cannot be achieved directly through it, and the final stages of the study of drug-receptor interactions should be evaluated before starting laboratory work by having effective compounds from the previous stages. Molecular dynamic simulations produce information at the microscopic level (position and velocity of atoms). The conversion of these data into macroscopic values (pressure, energy, etc.) is done using statistical mechanics. In fact, molecular dynamics and statistical mechanics link microscopic concepts and macroscopically observable quantities. Molecular dynamic simulations are only able to predict the thermodynamic behavior and stability of the ligand binding mechanism at the active site of the target enzyme. This is reported as a criterion.


#### 2.2.4. In Vitro

Study of drug in cell culture medium: effectiveness in these studies means inhibition of the replication of COVID-19 by the compound or drug under study [21].The half-maximal inhibitory concentrations (IC50) are a measure of the effectiveness of a compound in inhibiting biological function [22].In vivo studies are those in which the effects of drugs are tested on whole living organisms or cells usually animals as opposed to a tissue organism or dead organism. In vivo testing is better studied for observing the overall effects of an experiment on living subjects [23].


## 2.1. Data Sources and Searches

Studies published in PubMed, Scopus, Web of Science, EMBASE, Google Scholar, and DrugBank were searched from December 2019 to March 2021 using the following search terms: “Coronavirus,” “Covid-19,” “SARS-CoV-2,” “Drug design,” “Drug development,” “In silico,” “In vitro,” and “In vivo” alone or in combination without language restrictions. The keywords were selected using expert opinions, mesh, and related article titles.All articles with full text or in the absence of full text with abstract are included in the screening of this study. Studies were excluded if studies were comments, editorial, letters, review, and preprints.


## 2.2. Data Extraction

Two researchers independently extracted data from included studies using a predefined data extraction form. All disagreement was discussed and solved after rechecking the source data with a third investigator. The data extracted, including the last name of the first author, type of the study (in silico, in vitro, in vivo), and name of the agent (chemical compound, drug, herb, etc.), studied the mechanism and efficacy of the agent according to study design that specified according to the following definition.Computer-aided drug design can be divided into three different categories. All are based on ligands and receptors, which are briefly described [14] as follows:(1)
Dock Receptor-Based Approach [15]. Once the three-dimensional structure of the ligand molecules and their receptor is known, the receptor-based method is a good candidate for identifying or optimizing drugs. Due to the presence of three-dimensional structures of compounds and receptors, the nature of the interaction between the ligand and the receptor and the type of structure that the ligand can have to interact with them in favorable conditions can be identified using this method. The compound is simulated on the active site of the dock (meaning anchoring) and on the interaction of the ligand with the receptor by molecular mechanics and molecular dynamics. In this method, due to the ligation of the ligand in the active position, the ligand changes in terms of conformity and changes its position in different conditions and shows interaction with the receptor in different types of situations. To determine the type of ligands that can be docked into the receptor site, the matching of the shape and the complementarity of the hydrophobic, hydrophilic, and charged parts must be considered. Various software packages such as AUTODOCK-Glide-LUDI and LigandFit are used to design the drug based on the structure of the receptor.(2)
Ligand-Based Approach [16]. This method is used where the three-dimensional structures of the receptor are unknown and instead the structure of the ligands is known, which is one of the common methods. In this method, by indirectly studying compounds that react with biomolecules, they seek to design compounds that are pharmacologically active. In ligand-based drug design methods, in the absence of biomolecule structure, by studying specific ligands, it seeks to identify the structural and physicochemical properties of the compounds so that the desired compound can be designed based on data extracted from the study of previous compounds. This method is a kind of drug design based on pharmacophore (pharmacophore refers to the part of the drug to which the effect of the drug depends on that part of the molecule), and by studying the quantitative relationship between structure and their activity, drugs can be designed by this method. It can be said that it is a method for designing the pharmacophores of drugs.(3)
Denovo Design-Based Approach [17]. This method is used when the structure of the ligand is unknown but the structure of the receptor is known. In this method, there is information about the structures of the receptor or quasi-receptors, but there is no structure of the main composition that can interact with the active site of the receptor. One of the functions of drug design based on this method is to suggest and present the main composition that is complementary to the active site. The basis of the method is that the database of existing 3D structures is used to find small molecules that can interact with the active site of the receptor in terms of size, geometry, and functional groups. Software packages such as GROW and LEGEND are used to design drugs by this method.Drug design methods in the computer include quantitative structure-activity relationship (RASQ), docking, molecular dynamic simulation, and computational modeling. In these studies, the efficacy is evaluated based on the function of the drug or compound agent and the mechanism of action. Figure1 shows the methods of computer-aided drug design (CADD).Figure 1
Computational methods in drug design (CADD: computer-aided drug design).


### 2.2.1. Computational Methods in Drug Design

Quantitative Structure-Activity Relationship (QSAR). QSAR provides studies on the relationship between chemical structure and biological activity or other biological activities that are important in selecting or removing a compound before synthesis and testing. QSAR [18] is especially important to predict the result, especially when it is not possible to experiment with a compound. Molecular descriptors, which are the most important components of QSAR, can be obtained experimentally or through mathematical formulas from various theories such as quantum mechanics, chemical graph theory, and study theories. QSAR seeks to establish a statistically significant relationship between structure and performance. It also explains the specific effect of a drug and can ultimately predict the effect of newly synthesized chemical compounds. QSAR model is also an equation that predicts a property through molecular descriptors and their coefficients. Evaluation of the effectiveness of new compounds that have been studied using this method can be reported as a percentage of enzyme inhibition if the modeling has been done and mentioned in the article.


### 2.2.2. Docking

In this technique, to achieve a combination with a pharmacological effect and increase the pharmacological activity of the drug, different formulations of a drug interact with the receptor, and the structure that has the best interaction with the receptor and the lowest energy level is selected for laboratory steps [19]. In this way, possible structures that have a stronger interaction with the receiver can be isolated at this stage. The same issue is considered and reported as an effectiveness measure.


### 2.2.3. Molecular Dynamic Simulation

In this technique, which is based on the simulation of drug-receptor interaction in the body, docking problems are solved and in fact play a complementary role in this technique [20]. Due to the time-consuming work with this technique, effective structures cannot be achieved directly through it, and the final stages of the study of drug-receptor interactions should be evaluated before starting laboratory work by having effective compounds from the previous stages. Molecular dynamic simulations produce information at the microscopic level (position and velocity of atoms). The conversion of these data into macroscopic values (pressure, energy, etc.) is done using statistical mechanics. In fact, molecular dynamics and statistical mechanics link microscopic concepts and macroscopically observable quantities. Molecular dynamic simulations are only able to predict the thermodynamic behavior and stability of the ligand binding mechanism at the active site of the target enzyme. This is reported as a criterion.


### 2.2.4. In Vitro

Study of drug in cell culture medium: effectiveness in these studies means inhibition of the replication of COVID-19 by the compound or drug under study [21].The half-maximal inhibitory concentrations (IC50) are a measure of the effectiveness of a compound in inhibiting biological function [22].In vivo studies are those in which the effects of drugs are tested on whole living organisms or cells usually animals as opposed to a tissue organism or dead organism. In vivo testing is better studied for observing the overall effects of an experiment on living subjects [23].


## 2.2.1. Computational Methods in Drug Design

Quantitative Structure-Activity Relationship (QSAR). QSAR provides studies on the relationship between chemical structure and biological activity or other biological activities that are important in selecting or removing a compound before synthesis and testing. QSAR [18] is especially important to predict the result, especially when it is not possible to experiment with a compound. Molecular descriptors, which are the most important components of QSAR, can be obtained experimentally or through mathematical formulas from various theories such as quantum mechanics, chemical graph theory, and study theories. QSAR seeks to establish a statistically significant relationship between structure and performance. It also explains the specific effect of a drug and can ultimately predict the effect of newly synthesized chemical compounds. QSAR model is also an equation that predicts a property through molecular descriptors and their coefficients. Evaluation of the effectiveness of new compounds that have been studied using this method can be reported as a percentage of enzyme inhibition if the modeling has been done and mentioned in the article.


## 2.2.2. Docking

In this technique, to achieve a combination with a pharmacological effect and increase the pharmacological activity of the drug, different formulations of a drug interact with the receptor, and the structure that has the best interaction with the receptor and the lowest energy level is selected for laboratory steps [19]. In this way, possible structures that have a stronger interaction with the receiver can be isolated at this stage. The same issue is considered and reported as an effectiveness measure.


## 2.2.3. Molecular Dynamic Simulation

In this technique, which is based on the simulation of drug-receptor interaction in the body, docking problems are solved and in fact play a complementary role in this technique [20]. Due to the time-consuming work with this technique, effective structures cannot be achieved directly through it, and the final stages of the study of drug-receptor interactions should be evaluated before starting laboratory work by having effective compounds from the previous stages. Molecular dynamic simulations produce information at the microscopic level (position and velocity of atoms). The conversion of these data into macroscopic values (pressure, energy, etc.) is done using statistical mechanics. In fact, molecular dynamics and statistical mechanics link microscopic concepts and macroscopically observable quantities. Molecular dynamic simulations are only able to predict the thermodynamic behavior and stability of the ligand binding mechanism at the active site of the target enzyme. This is reported as a criterion.


## 2.2.4. In Vitro

Study of drug in cell culture medium: effectiveness in these studies means inhibition of the replication of COVID-19 by the compound or drug under study [21].The half-maximal inhibitory concentrations (IC50) are a measure of the effectiveness of a compound in inhibiting biological function [22].In vivo studies are those in which the effects of drugs are tested on whole living organisms or cells usually animals as opposed to a tissue organism or dead organism. In vivo testing is better studied for observing the overall effects of an experiment on living subjects [23].


## 3. Results

In this review, we reported a significant number of articles with in silico, in vitro, and in vivo approaches for drug development of COVID-19. We retrieved a total of 2538 articles from the initial database search. After the removal of duplication and screening, 317 studies were selected for inclusion in this review. Figure2 shows the PRISMA diagram.Figure 2
PRISMA diagram of the study.The analysis of article contents indicated that 266 studies performed in silico approaches against viral targets; 34 studies used in vitro approaches against SARS-CoV-2; and 15 studies used in vivo (animal) models.


### 3.1. Results from In Silico Drug Discovery

From 267 studies used in silico approaches, 98 article studies repurposed approved drugs with a new mechanism of action and 91 studies evaluated natural products (e.g., herbal medicine) on COVID-19. The characteristics of these studies are summarized in Tables1 and 2. Also, Table 3 shows the characteristics of the remaining studies (N = 87).Table 1
Summary of studies with in silico method that used repurposed drugs for novel drug discovery against COVID-19.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAbosheasha and El-Gowily [24]2020In silicoDrug repurposing molecular docking-based virtual screening15 antiplatelet FDA-approved drugsMain protease (Mpro) and spike glycoprotein (S)Cilostazol has favorable binding interaction with Mpro (PDB ID: 6LU7) cilostazol, iloprost, epoprostenol, prasugrel, and icosapent ethyl that have a higher binding affinity on spike glycoprotein (S)Cilostazol is a promising FDA drug against COVID-19 by inhibiting both Mpro and S proteinAbdul Kadhim et al. [25]2020In silicoDrug repurposing, dockingExperimental and approved drugsPapain-like protease and RNA polymeraseDrugs that shared >70% similarity to the binding sites of those targets were reversin, pentagastrin, remdesivir, norfloxacin, and nitazoxanide against COVID-19 papain-like protease whereas benzylglutathione, lopinavir, and hydroxymethylglutathione against RNA polymeraseAntiresistance reversin showed the highest inhibitory efficacy against COVID-19 papain-like protease, and benzylglutathione is an experimental compound; however, it had the highest RNA polymerase inhibiting efficacyAbu-Saleh et al. [26]2020In silicoLigand-based/structure-based virtual screening, MD simulations, and binding energy calculationsApproved drugs and bioactive compounds listed in the DrugBank and ChEMBL databasesMain proteaseBest MM/GBSA binding energy; ChEMBL275592, montelukast, ChEMBL28834. Bromocriptine and saquinavir demonstrate stability in the active site of MproAchilonu et al. [27]2020In silicoHigh-throughput virtual screening and ligand dockingFDA-approved drugsMain proteaseIsavuconazonium, a P2-P3α-ketoamide derivative, and pentagastrina are the top three molecules (Lig13b as the benchmark) based on docking energyAftab et al. [28]2020In silicoRepositioning/target-based virtual screening and molecular dockingTen antiviral drugs were screened: ribavirin, remdesivir, sofosbuvir, penciclovir, nitazoxanide, nafamostat, chloroquine, galidesivir, favipiravir, and interferonRNA-dependent RNA polymerase (RdRp)Galidesivir and its drug-like compounds CID123624208 and CID11687749 have shown an effective attachment to the priming site of viral RdRpCID123624208 and CID11687749 may be considered for in vitro and in vivo clinical trialsAhmadi et al. [29]2021In silicoDrug repurposing study using molecular dockingEnfuvirtide, an HIV-1 fusion inhibitor peptideSARS-CoV-2 fusion inhibitorEnfuvirtide binding to the S2 protein of SARS-CoV-2 was remarkably stable and can act as a strong SARS-CoV-2 fusion inhibitorAhmed et al. [30]2021In silicoDrug repurposing (high-throughput virtual screening) (HTVS) followed by re-docking with standard precision (SP) and extra-precision (XP) molecular dockingFDA-approved antiviral and anti-infection drugsMain proteaseOf 1397 potential drugs, 157 showed considerable affinity towards Mpro. High-affinity lead drugs (iodixanol, amikacin, troxerutin, and rutin) were identified. Amikacin was found to be the most potent inhibitor of main proteaseAminoglycosides may serve as a scaffold to design potent drug molecules against COVID-19Anand et al. [31]2021In silicoMolecular docking130 US FDA-approved drugs including hypertension, cardiovascular diseases, respiratory tract infections (RTI), antibiotics, and antiviral drugsStructural and nonstructural proteins of SARS-CoV-2 (nsp3, nsp5, nsp10, nsp16)15 potent drugs exhibiting significant inhibitory potential against SARS-CoV-2 like baloxavir marboxil, danoprevir and sofosbuvir, fosinopril, moexipril, quinapril, telmisartan, azilsartan, verapamil, and doxazosinAzithromycin, doxycycline, clarithromycin, rifamycin, and augmentin: nsp10; virginiamycin, tunicamycin, quinupristin, fidaxomicin, digoxin, and azithromycin: main protease; caspofungin, amphotericin B, ketoconazole, and micafungin: E and N proteins; virginiamycin and amphotericin B: S proteinAncy et al. [32]2020In silicoMolecular docking, molecular dynamics, and binding free energy simulation studyHIV-1 protease, namely, TMB607 and TMC-310911Main proteaseTMB607 molecule binds strongly with the SARS-CoV-2 main protease enzymeAnsari et al. [33]2020In silicoRepurposing drug molecular dockingTAT-peptide 47–57 (GRKKRRQRRRP)-conjugated repurposed drugs (i.e., lopinavir, ritonavir, favipiravir, and hydroxychloroquine)Main proteaseTP-conjugated ritonavir, lopinavir, favipiravir, and hydroxychloroquine have superior and significantly enhanced interactions with main proteaseArun et al. [34]2020In silicoRepurposing drug molecular dockingDrugs available in the super DRUG2 databaseMain proteaseBinifibrate and bamifylline bind strongly to the enzyme active siteArya et al. [35]2020In silicoMolecular dockingFDA-approved drugsPapain-like protease15 FDA-approved drugs, including chloroquine and formoterol, bind the target enzyme with significant affinity and good geometry, suggesting their potential to be utilized against the virusBaby et al. [36]2020In silicoSchrodinger’s computer-aided drug discovery tools for in silico drug repurposingFDA-approved library of drugsRNA-dependent RNA polymerase (RdRp)Pitavastatin, ridogrel, rosoxacinBaby et al. [36]2021In silicoSchrodinger’s computer-aided drug discovery tools for in silico drug repurposingFDA-approved library of drugsMain proteaseTipiracil and aprepitant interacted with the main proteaseBaker et al. [37]2021In silicoRepurposing drug molecular docking50 compounds with activity against main proteaseMain proteaseDrugs including boceprevir, ciluprevir. narlaprevir, and telaprevir may be more potent against main protease than boceprevir and suitable for rapid repurposingBharath et al. [38]2020In silicoDrug repurposing computer-aided drug design (CADD)4015 known and approved small moleculesSpike glycoproteinGlycyrrhizic acid (GA) of plant origin may be repurposed for SARS-CoV-2 interventionBhowmik et al. [39]2021In silicoRepurposing drugs, docking, and molecular dynamic simulationOrientin (phytochemical)Inhibitor of SARS-CoV-2 spike and host cell receptor GRP78 bindingBinding of orientin in the overlapping residues of GRP78 binding region of SARS-CoV-2 spike modelAs a promising precautionary or therapeutic measure for COVID-19Bolelli et al. [40]2021In silicoDrug repurposing, virtual screening methodFDA-approved drugsMain proteaseThree compounds (dobutamine and its two derivatives)Cavasotto et al. [41]2021In silicoDrug repurposing, docking-based screening using a quantum mechanical scoringFDA-approved drugsSpike protein, main protease papain-like proteaseSovaprevir, elbasvir, danoprevir, samatasvir, Candesartan, saquinavir ritonavir, indinavir, lopinavir, brilacidin, flovagatran, aplidin, desmopressin, and felypressin listed as potential inhibitors of main proteaseChandel et al. [42]2020In silicoDrug repurposing, molecular dynamics, and dockingFDA-approved drugsNsp9 replicase and spike proteinsConivaptan exhibited the highest binding of the Nsp9 replicase. Tegobuvir exhibited maximum stability along with the highest binding energy at the active site of the spike proteinsChen et al. [43]2020In silicoDrug repurposing, molecular dynamics, and dockingFDA-approved drugsSpike (S)-mediated cell entryCepharanthine, abemaciclib, osimertinib, trimipramine, colforsin, and ingenolChidambaram et al. [44]2020In silicoMolecular dockingCoumarins and their analogsMain proteaseNatural coumarin analog toddacoumaquinone displayed remarkable inhibition ability. Synthetic coumarin analog (1 m) also displayed the comparable inhibition ability main protease in intricate withα-ketoamideChoudhary et al. [45]2020In silicoDrug repurposing, molecular dynamics, and dockingFDA-approved drugsSpike glycoprotein and cellular angiotensin-converting enzyme 2 (ACE2) receptorGR 127935 hydrochloride hydrate, GNF-5, RS504393, TNP, and eptifibatide acetate were found binding to virus binding motifs of ACE2 receptor. KT203, BMS195614, KT185, RS504393, and GSK1838705A were identified to bind at the receptor-binding site on the viral S proteinClemente et al. [46]2021In silicoMolecular docking, molecular dynamic (MD) simulationsIbuprofenMain proteaseRacemic mixtures of the ibuprofen enantiomers might be a potential treatment for main proteaseCosic et al. [47]2021In silicoExtended resonant recognition model (RRM)IvermectinSpike proteinsIvermectin could interfere with activity of spike proteinsDaoud et al. [48]2021In silicoStructure-based pharmacophore approach, molecular docking, and repurposing studiesFDA-approved drugsMain proteaseLopinavir, remdesivir, ritonavir, saquinavir, and raltegravir were successfully docked into the binding site of main proteasede Oliveira et al. [49]2021In silicoMolecular modeling and virtual screening and repurposing studies9091 FDA-approved drugsSpike protein24 best scored ligands (14 traditional herbal isolates and 10 approved drugs) as potential candidates to inhibit the S proteinQuinupristin, nilotinib, acetyldigitoxinDelijewski and Haneczok [50]2021In silicoSupervised machine learning model and repurposing studiesFDA-approved drugsAgainst SARS-CoV-2Zafirlukast as the best repurposing candidate for COVID-19Dey et al. [51]2021In silicoVirtual database screening, molecular docking, all-atom molecular dynamic simulation, and MM-PBSA analysisTretinoin, mefenamic acid, ondansetron, and artemetherEnvelope (E) proteinTretinoin as a potential SARS-CoV-2 E protein ion channel blocker and virus assembly inhibitorDurdagi [52]2020In silicoMolecular modeling approach in virtual drug screening repurposing studyFDA-approved drugsType 2 transmembrane serine protease (TMPRSS2)Benzquercin as strong TMPRSS2 inhibitorDurdagi et al. [53]2020In silicoMolecular docking, MM-GBSA-based predictions, and molecular dynamic repurposing studyFDA-approved drugsMain protease and spike receptor-binding domain bound with ACE2 COVID-19 target proteinsPimelautide, rotigaptide, telinavir, ritonavir, pinokalant, terlakiren, cefotiam, and cefpiramide as SARS-CoV-2 main protease inhibitors. Denopamine, bometolol, naminterol, rotigaptide, and benzquercin as potential ACE2/spike protein domain inhibitorsEleftheriou et al. [54]2020In silicoMolecular docking34 approved and on-trial protease inhibitorsMain proteaseHCV protease, DPP-4,α-thrombin, and coagulation factor Xa known inhibitorsElmezayen et al. [55]2021In silicoMolecular modeling approach in virtual drug screening repurposing studyCommercially available drugs and ZINC15 libraryMain proteasesFour potential inhibitors against Mpro enzyme, two available drugs (Talampicillin and Lurasidone), and two novel drug-like compounds (ZINC000000702323 and ZINC000012481889)Encinar et al. [56]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study9000 US Food and Drug Administration (FDA)-approved investigational and experimental drugs from the DrugBank repositoryS-Adenosyl-L-methionine-binding pocket of nsp16, [2] the unique “activating surface” between nsp16 and nsp10, and [3] the RNA-binding groove of nsp16Tegobuvir, sonidegib, siramesine, antrafenine, bemcentinib, itacitinib, or phthalocyanine antagonism of SARS-CoV-2 RNAs lacking 20-O-methylationFarag et al. [57]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study2000 FDA-approved drugsMain proteaseDarunavir, nelfinavir, and saquinavir bound to the central site of main protease substrate-binding pocket rosuvastatin, montelukast, and the anti-histaminic fexofenadine bound to the terminal site of main protease substrate-binding pocketStarting point for further in vitro and in vivo testingFeng et al. [58]2020In silicoMolecular modeling approach in virtual drug screening and repurposing studyFDA-approved drugsSpike proteinEltrombopag possesses a high binding affinity to S protein plus human ACE2Ferraz et al. [59]2020In silicoLigand and structure-based virtual screening, repurposing studyFDA-approved drugsMain proteaseTwo oral (bedaquiline and glibenclamide) and one buccal drug (miconazole)Fischer et al. [60]2020In silicoMolecular docking approach in virtual drug screening and repurposing studyOver 606 million compoundsMain protease12 purchasable compounds, with binding affinity to the target protease the natural compounds (−)—taxifolin and rhamnetin as potential inhibitors of main proteaseGimeno et al. [61]2020In silicoMolecular modeling approach in virtual drug screening and repurposing studyFDA-approved drugsMain proteasePerampanel, carprofen, celecoxib, alprazolam, trovafloxacin, sarafloxacin, and ethyl biscoumacetate. Carprofen and celecoxibInitiative for in vitro testingGuo et al. [62]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study single-cell RNA sequencingUS FDA-approved drugsAgainst SARS-CoV-2281 FDA-approved drugs that have the potential to be effective against SARS-CoV-2 infection, 16 of which are currently undergoing clinical trials to evaluate their efficacy against COVID-19Including the HIV protease inhibitor lopinavir/ritonavir combination (phase 4), glucocorticoid receptor agonist dexamethasone (phase 3/4), DNA replication inhibitor niclosamide (phase 2/3), antineoplastic agent lenalidomide (phase 4), and calcineurin inhibitor tacrolimus (phase 3), ABT-737 (BCL inhibitor), brefeldin-A (protein synthesis inhibitor), indirubin (CDK inhibitor), TPCA-1 (IKK inhibitor), lopinavir (HIV protease inhibitor), GW-441756 (growth factor receptor inhibitor), treprostinil (prostacyclin analog), tyrphostin-AG-1478 (EGFR inhibitor) and epoxycholesterol (LXR agonist), fostamatinib (SYK inhibitor), VER-155008 (HSP inhibitor), KU-0063794 (MTOR inhibitor), PIK-90 (PI3K inhibitor), linsitinib (IGF-1 inhibitor), TAK-715 (p38 MAPK inhibitor), Y-27632 (Rho-associated kinase inhibitor), AZ-628 (RAF inhibitor), and lestaurtinib (FLT3 inhibitor)Gupta et al. [63]2020In silicoMolecular modeling approach in virtual drug screening and repurposing studyFDA-approved drugsMain proteaseCobicistat is the most efficient inhibitor of Mpro both in silico and in vitroHuynh et al. [64]2021In silicoDocking and molecular dynamics and repurposing studyFDA-approved drugsPapain-like proteaseThe chances of drug repurposing for PLpro might be lowIbrahim et al. [65]2020In silicoMolecular dynamic simulations, molecular docking, MM-GBSA analysis, and repurposing studyDrugBank databaseMain proteaseDB02388 and cobicistat (DB09065)Iftikhar et al. [66]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study4574 compounds also containing FDA-approved drugsRdRp, main protease, and helicaseRimantadine, bagrosin, and grazoprevir showed binding to main protease. Casopitant is a neurokinin-1 receptor that showed binding to RdRp. Meclonazepam and oxiphenisatin showed specific interactions with helicaseJain and Mujwar [67]2020In silicoComputational drug repurposing docking simulations2880 FDA-approved drugsMain proteaseMetocurine, dihydroergotoxine, imatinib, daunorubicin, bromocriptine, irinotecan, azelastine, gestodene, adapalene, and simvastatinMetocurine was chosen as a safe and effective drug candidate for developing therapy against the viral Mpro enzyme of SARS-CoV-2 for the treatment of COVID-19Jarvis et al. [68]2020In silicoTier-based scoring system repurposing studyClinically developed drugsPotential repurposing against COVID-19Four drug classes (antimalarial amino-quinolones, selective estrogen receptor modulators (SERMs), low potency tricyclic antipsychotics, and tricyclic antidepressants) as potential drug candidates for COVID-19The tricyclic antipsychotics and tricyclic antidepressants were further excluded based on a high adverse event profileKadioglu et al. [69]2021In silicoRepositioning/virtual drug screening, molecular docking, and supervised machine learning algorithm drug repositioningFDA-approved drug natural compound dataset ZINC databaseSpike protein, nucleocapsid protein, and 2′-o-ribose methyltransferaseConivaptan, paritaprevir, simeprevir, dihydroergotamine, ZINC000027215482, ZINC000252515584, loniflavone, procyanidinKandeel et al. [70]2020In silicoDrug repurposing molecular dynamic (MD) simulations followed by molecular mechanics/generalized born surface area (MM/GBSA) binding energy calculations1697 clinical FDA-approved drugsPapain-like proteasePhenformin, quercetin, and ritonavirPhenformin was more stable than quercetin and ritonavirKandwal and Fayne [71]2020In silicoRepurposing drug computational design pharmacophore featuresIn-development/approved drugsViral nucleocapsid and nonstructural proteinsIsepamicin and streptomycin (nsp3); coenzyme-I, rutin, epigallocatechin gallate-(-), and procyanidin-b-2 (nsp7/nsp8/nsp12); paromomycin (nsp10/nsp16); olomoucine, sapropterin, tetrahydrofolic acid, INS316, and adenosine phosphate (nsp15); varespladib, hexanoic acid, citric acid, OSI-027, MK-5108, stepronin, calcium gluceptate, CPP, pirenoxine, midafotel, and maltobionic acid (nucleocapsid)Khan et al. [72]2020In silicoDrugs repurposing molecular dynamic simulation31 FDA-approved anti-HIV drugs, and traditional Chinese medicines (TCM) databaseMain proteaseSaquinavir and TCM5280805Khan et al. [73]2020In silicoDrugs repurposing molecular docking23 prospective drug candidatesMain proteaseEpirubicin, vapreotide, and saquinavir exhibited better binding affinitySynergistic interactionKouznetsova et al. [74]2020In silicoDrugs repurposing molecular dockingFDA-approved drugsPapain-like proteaseInhibitors of HIV, hepatitis C, and cytomegalovirus (CMV) demonstrated some activityKrishnaprasad et al. [75]2020In silicoDrugs repurposing molecular dockingFDA-approved library of drugsRNA-dependent RNA polymerasePitavastatin, ridogrel, and rosoxacin displayed superior binding with the active siteKumar et al. [76]2020In silicoDrugs repurposing docking and molecular dynamic (MD) simulations combined with molecular mechanics/generalized born surface area (MM/GBSA)12 FDA-approved drugs (darunavir, indinavir, saquinavir, tipranavir, diosmin, hesperidin, rutin, raltegravir, velpatasvir, ledipasvir, rosuvastatin, and bortezomib)Main proteaseSaquinavir as a potent inhibitor of dimeric main proteaseKumar et al. [77]2020In silicoDrugs repurposing molecular docking molecular dynamic simulations MM/GBSAWithaferin A (Wi-A), withanone (Wi-N) (active withanolides of ashwagandha), and caffeic acid phenethyl ester (CAPE, bioactive ingredient of propolis)Main proteaseWi-N and CAPE possess the potential to inhibit the functional activity of main proteaseKumar et al. [78]2020In silicoDrugs repurposing molecular dockingFDA-approved drugsMain proteaseLopinavir-ritonavir, tipranavir, and raltegravir show the best molecular interaction with the main proteaseKumar et al. [79]2020In silicoDrugs repurposing molecular docking molecular dynamic simulationsFDA-approved library of drugsMain proteaseHyaluronic acid and acarbose show strong interactions with catalytic site residues of main proteaseLi et al. [80]2020In silicoDrug repurposing free energy perturbation-based absolute binding free energy (FEP-ABFE) predictionsVirtual screening of existing drugsMain protease25 drugs were predicted, and 15 were confirmed as potent inhibitors of SARS-CoV-2 main protease. The most potent one is dipyridamole. Hydroxychloroquine (ki = 0.36μM) and chloroquine (ki = 0.56 μM) were also found to potently inhibit main proteaseLiang et al. [81]2021In silicoDrug repurposing molecular docking2,631 FDA-approved small moleculesMultiple main proteins29 drugs that could actively interact with two or more target proteins, with 5 drugs (avapritinib, bictegravir, ziprasidone, capmatinib, and pexidartinib) being common candidates for all four key host proteins and 3 of them possessing the desirable molecular propertiesLokhande et al. [82]2021In silicoDrugs repurposing molecular docking molecular dynamic simulationsFDA-approved drugsMain proteaseMitoxantrone, leucovorin, birinapant, and dynasoreMahanta et al. [83]2020In silicoDrugs repurposing molecular docking molecular dynamic simulationsU.S. Food and Drug Administration-approved antimicrobial drugsMain proteaseViomycinMahdian et al. [84]2021In silicoDrugs repurposing molecular docking molecular dynamic simulationsFDA-approved drugsViral entry receptors (ACE2 and CD147) and integral enzyme of the viral polymerase (RdRp)Ledipasvir, estradiol benzoate, and vancomycin and paritaprevirMarak et al. [85]2020In silicoRepurposing drug homology modeling molecular docking108 FDA-approved antiparasitic and anti-inflammatory drugs10 SARS-CoV-2 targets (PLpro, 3CLpro, RdRp, spike, helicase, NSP1, NSP3, NSP4, NSP9, and NSP16-NSP10)Ivermectin, atovaquone, posaconazole, doxycycline, moxidectin, amphotericin B, chlortetracycline, spiramycin, sulfasalazine, parecoxib, and etoricoxib exhibited good binding affinitiesMohapatra et al. [86]2020In silicoRepurposing drug machine learning (ML) technologyFDA-approved drugsAgainst COVID-1910 FDA-approved commercial drugs that can be used for repurposing amprenavir would probably be the most effective drug based on the selected criteriaMolavi et al. [87]2021In silicoRepurposing drug molecular docking1760 FDA-approved drugsRNA-dependent RNA polymerase (RdRp) and main proteaseNilotinib, imatinib, and dihydroergotamine for 3clpro and dexasone and raltegravir for RdRp. Raltegravir, an anti-HIV drug, was observed to be the best compound against RdRp based on docking binding energy dihydroergotamine is a suitable candidate for main proteaseMulgaonkar et al. [88]2020In silico/in vitroRepurposing drug molecular dockingFDA-approved drugsSpike glycoproteinBCR-ABL tyrosine kinase inhibitor, imatinib, inhibits SARS-CoV-2Via fusion inhibitionMycroft-West et al. [89]2020In silicoRepurposing drug molecular docking molecular dynamic simulationsHeparinSpike (S1) protein receptor-binding domainInhibition of viral infection arises from an overlap between the binding sites of heparin/HS on S1-RBDRepurposing heparin and its derivatives as antiviral agents against SARS-CoV-2Nayarisseri et al. [90]2020In silicoShape-based machine learning assisted by molecular docking and molecular dynamic simulations. ADMET studies31 repurposed compoundsMain proteaseRemdesivir, valrubicin, aprepitant, and fulvestrantThe novel compound nCorv-EMBS herein proposed stands as a promising inhibitor to be evaluated further for COVID-19 treatmentOdhar et al. [91]2020In silicoMolecular docking molecular dynamic simulations1615 FDA-approved drugsMain proteaseConivaptan azelastineOrtega et al. [92]2020In silicoRepurposing drug molecular dockingFamotidineAgainst SARS-CoV2Famotidine could interact within the catalytic site of the three proteases associated with SARS-CoV2 replicationWeak binding affinity could be reached only upon intravenous administrationPandey et al. [93]2021In silicoRepurposing drug molecular docking9 flavonoidsSpike glycoproteinBaicalinParveen and Alnoman [94]2021In silicoMolecular docking molecular dynamic simulation density functional theory (DFT) ADME-ToxFDA-approved anticancer drugs (capmatinib, pemigatinib, selpercatinib, and tucatinib)Spike glycoprotein (S1) and the main proteasePotential of selected anticancer drugs for plausible drug development to fight COVID-19Capmatinib, pemigatinib, selpercatinib, and tucatinibPeele et al. [95]2020In silicoMolecular docking molecular dynamic simulationsUSFDA-approved drugs, plant-derived natural drugsMain proteaseLopinavir, amodiaquine, theaflavin digallatePinzi et al. [96]2021In silicoDrug repurposing molecular docking molecular mechanic Poisson–Boltzmann surface area (MM-PBSA)DrugBank databaseMain protease22 candidates with putative SARS-CoV-2 Mpro inhibitory activity. Enalkiren, ethylsulfonamide-D-Trp-Gln-p-aminobenzamidine, delparantag ritonavir and lopinavir, saquinavirBeneficial polypharmacological effectsPokhrel et al. [97]2020In silicoDrug repurposing molecular dynamic simulationsUS Food and Drug Administration (FDA)-approved drugsRNA-dependent RNA polymeraseQuinupristin is particularly interesting because it is expected to bind across the RNA tunnel, blocking access from both sidesQuinupristin represents a potential anti-SARS-CoV-2 therapeuticRay et al. [98]2020In silicoDrug repurposing intramolecularly quenched fluorescence (IQF) peptide substrate774 FDA-approved drugsMain proteaseEthacrynic acid, naproxen, allopurinol, butenafine hydrochloride, raloxifene hydrochloride, tranylcypromine hydrochloride, saquinavir mesylateSachdeva et al. [99]2020In silicoDrug repurposing molecular dockingAntimalarial drugsSpike protein and main proteaseDoxycycline showed the most effective binding to the spike protein, whereas halofantrine and mefloquine bound effectively with the main proteaseDoxycycline could potentially be a good candidate for repurposing for COVID-19Sang et al. [100]2020In silicoDrug repurposing molecular docking molecular mechanics Poisson–Boltzmann surface area (MM-PBSA)6 approved anti-HIV drugsMain proteaseDarunavirSaxena et al. [101]2021In silico/In vitroDrug repurposing molecular dockingFDA-approved DrugBank databaseSpike proteinErtugliflozin possesses several desired propertiesGood candidate for immediate repurposing for the treatment of COVID-19Setianingsih et al. [102]2020In silicoDrug repurposing molecular docking, molecular dynamic simulations160 potential drugs from therapeutic target database13 protein targets (12 SARS-CoV-2 proteins and 1 human protein)Suramin, the strongest binding affinity against 3 protein targets (spike protein, nucleocapsid protein, ACE2)Suramin is the most potential to bind nucleocapsid and spike protein of SARS-CoV-2Shah et al. [103]2020In silicoDrug repurposing molecular docking61 molecules that are already being used in clinics or under clinical scrutiny as antiviral agentsAgainst the SARS-CoV-237 molecules were found to interact with >2 protein structures of COVID-19. HIV protease inhibitors and RNA-dependent RNA polymerase inhibitors showed promising features of binding to COVID-19 enzymeMethisazone, an inhibitor of protein synthesis; CGP42112A, an angiotensin AT2 receptor agonist; and ABT450, an inhibitor of the nonstructural protein 3-4A, might become convenient treatment option as well against COVID-19Sharma and Mishra [104]2020In silicoDrug repurposing target-based virtual ligand screeningZINC drug database and our own database of natural productsAgainst the SARS-CoV-2Antivirus drugs (ribavirin, valganciclovir, and thymidine), antibacterial drugs (cefpiramide, sulfasalazine, phenethicillin, lymecycline, demeclocycline, doxycycline, oxytetracycline, and tigecycline), anti-asthmatic drugs (montelukast, fenoterol, and reproterol), and hepatoprotective drug silybin have antiviral activity. Natural hesperidin was targeting the binding between spike RBD and human ACE2The natural products, such as flavonoids like neohesperidin, hesperidin, baicalin, kaempferol 3-O-rutinoside, and rutin from different sources, andrographolide, neoandrographolide, and 14-deoxy-11,12-didehydroandrographolide from A. paniculata, and a series of xanthones from the plants of Swertia genus, with antivirus, antibacteria, and anti-inflammation activity could effectively interact with these targets of SARS-CoV-2Shekhar et al. [105]2020In silicoDrug repurposing molecular docking molecular dynamic simulations2,625 FDA-approved small moleculesSpike (S) protein fusion peptide regionChloramphenicol succinate, imipenem, and imidureaSingh et al. [106]2021In silicoDrug repurposing molecular docking molecular dynamic simulations1749 FDA-approved drugsNSP12, a RNA polymerase5 compounds which include 3a (paritaprevir), 3d (glecaprevir), 3h (velpatasvir), 3j (remdesivir), and 3l (ribavirin) had the best binding affinitySinha et al. [107]2021In silicoDrug repurposing systematic pharmacokinetics, drug-likeness, basicity predictions, virtual screening, and molecular dynamic analysisHydroxychloroquine (HCQ), chloroquine (CQ)Spike protein1-[1-(6-Chloroquinolin-4-yl) piperidin-4-yl]piperidin-3-ol and (1r,2R)-2-N-(7-chloroquinolin-4-yl)cyclohexane-1,2-diamine interact with the active site of the spike protein similar to HCQ and CQ, respectively, with augmented safety profileSoni et al. [108]2020In silicoMolecular docking molecular dynamic simulation ADME propertiesRifampicinMain proteaseRifampicin docking score was −7.24 kcal·mol–1, and it can predict as a very good inhibitor of main proteaseTariq et al. [109]2020In silicoDrug repurposing molecular docking molecular dynamic simulations15 antimalarial drugs (including chloroquine) and 2413 US Food and Drug Administration-approved drugsMain protease spike (S) proteinParomomycin with activity against two targets spike protein and protease domainTatar et al. [110]2021In silicoDrug repurposing molecular docking molecular dynamic simulations34 antiviral compoundsRNA-binding domainRapamycin, saracatinib, camostat, trametinib, and nafamostat were the top hit compoundsTejera et al. [111]2020In silicoDrug repurposing quantitative structure-activity relationship (QSAR) mode molecular docking molecular dynamic simulation MM-PBSA methodDrugBank databaseMain proteaseLevothyroxine, amobarbital, and ABP-700Teralı et al. [112]2020In silicoDrug repurposing molecular docking7,173 clinically approved drugAngiotensin-converting enzyme 2 (ACE2)Lividomycin, burixafor, quisinostat, fluprofylline, pemetrexed, spirofylline, edotecarin, diniprofyllineTrezza et al. [113]2020In silicoDrug repurposing docking simulations, with molecular dynamics (MD), supervised MD (SuMD), and steered MD (SMD) simulationsFDA-approved drugsSpike glycoproteinSimeprevir, lumacaftorUgurel et al. [114]2020In silicoDrug repurposing structure-based drug design genome sequences were analyzedFDA-approved drugsHelicase (Nsp13)Cangrelor, fludarabine, folic acid, and polydatin inhibit both the wild-type and mutant SARS-CoV-2 helicaseUnni et al. [115]2020In silicoDrug repurposing molecular docking molecular dynamic simulationsDrugBank and PubChem librarySpike protein (S protein)Bisoxatin (DB09219)A laxative drugVaishali et al. [116]2020In silicoDrug repurposing molecular docking molecular dynamic simulation ADME propertiesFDA-approved compoundsNonstructural protein 9 (Nsp9) replicase and spike proteinsConivaptan exhibited the highest binding energy and maximum stability of the Nsp9 replicase. Tegobuvir exhibited maximum stability along with the highest binding energy at the active site of the spike proteinsVerma et al. [117]2020In silicoDrug repurposing molecular docking molecular dynamic simulation MM-GBSA-based energyFDA-approved drugsMain proteaseTop-ranked drugs including adefovir, lumefantrine, dipyridamole, dihydroergotamine, hexoprenaline, riboflavin (vitamin B2), and pantethine (vitamin B5)Wei et al. [118]2020In silicoDrug repurposing molecular docking molecular dynamic simulationsUS Food and Drug Administration (FDA)-approved drugs from DrugBank and natural compounds from traditional Chinese medicine systems pharmacology (TCMSP)Spike protein (S protein)Digitoxin and bisindigotin in TCMSP had the highest docking scores Forsythiae fructus and Isatidis radix are components of Lianhua Qingwen, and raltegravir had relatively high binding scoresXu et al. [119]2021In silicoDrug repurposing molecular docking molecular dynamic simulationsFDA-approved drugsSpike proteinThymoquinone, a phytochemical compound obtained from the plant Nigella sativa, is a potential drug candidateTable 2
Summary of studies with in silico method that used natural products for novel drug discovery against COVID-19.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAbdelli et al. [120]2021In silicoMolecular dockingIsothymol, thymol, limonene, P-cymene, andγ-terpinene derived from the essential oil of the antiviral and antimicrobial plant Ammoides verticillata (Desf.) Briq.Inhibition of ACE2 cellular receptorIsothymol, a major component of this plant, gives the best docking scores, as good ACE2 inhibitorAbouelela et al. [121]2021In silicoMolecular docking, dynamic simulation, and binding free energy calculationAloeMain protease and spike protein132, 134, and 159 were the best scoring compounds against main protease, while compounds 115, 120, and 131 were the best scoring ones against spike glycoprotein. Compounds 120 and 131 were able to achieve significant stability and binding free energies during molecular dynamic simulationAdem et al. [122]2020In silicoMolecular dockingMedicinal plant-based bioactive compounds (80 flavonoid compounds)Main proteaseHesperidin, rutin, diosmin, apiin, diacetylcurcumin, (E)-1-(2-hydroxy-4-methoxyphenyl)-3-[3-[(E)-3-(2-hydroxy-4-methoxyphenyl)-3-oxoprop-1-enyl]phenyl]prop-2-en-1-one, and beta, beta′-(4-methoxy-1,3-phenylene)bis(2′-hydroxy-4′,6′-dimethoxyacrylophenone have been found as more effective on COVID-19 than nelfinavirAllam et al. [123]2020In silicoMolecular docking, 3D shape similarity study (rapid overlay chemical similarity-ROCS) to the clinically used drugs in COVID-19 patients3′-Hydroxy-4′-methoxy-chroman-7-O-β-d-glucopyranoside 4, ferulic acid heptyl ester 1, naringenin 2, and 4,2′,4′-trihydroxy-6′-methoxychalcone-4′-O-β-d-glucopyranoside 3, which were isolated from peach (Prunus persica (L.) Batsch) fruitsMain protease, spike proteinNaringenin 2 and 4,2′,4′-trihydroxy-6′-methoxychalcone-4′-O-β-d-glucopyranoside 3 have a strong binding mode to a protease receptor and spike protein and also block the inflammatory stormRecommendation of peach fruits in controlling and managing COVID-19 casesAl‐Sehemi et al. [124]2020In silicoMolecular docking31000 natural compounds of the natural product activity and species source (NPASS) librarySpike glycoproteinCastanospermine and karuquinone B were shown to be the best-in-class derivatives in silico able to target an essential structure of the virus and to act in the early stage of infectionAttia et al. [125]2021In silicoMolecular docking10 phenolic antiviralAgainst SARS-CoV-2Hesperidin showed the highest docking scoreHesperidin and its mediated ZnO nanoparticles are willing antiviral agentsAzim et al. [126]2020In silicoVirtual screening methods molecular docking27 plant metabolitesMain protease proteins (MPP), Nsp9 RNA-binding protein, spike receptor-binding domain, spike ectodomain, and HR2 domainAsiatic acid, avicularin, guajaverin, and withaferin showed a maximum binding affinity with all key proteins in terms of lowest global binding energyBhowmik et al. [39]2021In silicoRepurposing drugs, docking, and molecular dynamic simulationOrientin (phytochemical)Inhibitor of SARS-CoV-2 spike and host cell receptor GRP78 bindingBinding of orientin in the overlapping residues of GRP78 binding region of SARS-CoV-2 spike modelAs a promising precautionary or therapeutic measure for COVID-19Çakır et al. [127]2021In silicoMolecular dockingPeptides derived from beta-lactoglobulinInhibit the host cell membrane receptorsAla-Leu-Pro-Met-His-Ile-Arg (ALMPHIR) and Ile-Pro-Ala-Val-Phe-Lys (IPAVFK) peptidesβ-Lactoglobulin (BLG) is the major whey protein of cow and sheep’s milk (∼3 g/l)Chatterjee et al. [128]2021In silicoMolecular dockingHesperidin, kaempferol, quercetin, epigallocatechinPLpro (papain-like protease), RdRp (RNA-dependent RNA polymerase), Mpro or 3cl protease, and spike proteinHesperidin, kaempferol, quercetin, epigallocatechinLead to conclusive data for the treatment of polyphenols, flavonoids, and bioflavonoids against SARS-CoV-2Chikhale et al. [129]2020In silicoMolecular docking, dynamicsPlant Withania somnifera (Indian ginseng)NSP15 endoribonuclease and receptor-binding domain of prefusion spike proteinWithanoside X and quercetin glucoside from W. somnifera have favorable interactions at the binding site of selected proteinsImmunomodulatory, antioxidant, and anti-inflammatory rolesChikhale et al. [130]2021In silicoMolecular docking, dynamics, and network pharmacology analysisSaikosaponinsAdjuvant therapy in the treatment of COVID-19Saikosaponins interact with the proteins CAT gene CAT (catalase) and checkpoint kinase 1 (CHEK1)Possible improvement in immune response towards COVID-19Chikhalet al. [131]2020In silicoMolecular docking, dynamicsAsparagus racemosus (Willd.)NSP15 endoribonuclease and spike receptor-binding domainAsparoside -C and Asparoside -F have good binding with target proteinsAsparagus racemosus holds promise as SARS-CoV-2 (S) and (N) protein inhibitorChowdhury [132]2020In silicoMolecular docking, dynamicsTinospora cordifolia (Giloy)Main proteaseBerberine can regulate main protease protein’s functionDahab et al. [133]2020In silicoMolecular docking10 phenolic compounds of different classes (phenolic acids, flavonoids, and coumarins)Main protease and RNA polymeraseThe top 7 hits are flacourticin [3], sagerinic acid [16], hordatine a [23], hordatine B [134], N-feruloyl tyramine dimer [135], bisavenanthramides B-5 [27], and vulnibactins [38] and have better binding scores than remdesivir, the native ligand in RNA polymerase target (PDB ID: 7bV2)Hordatines are phenolic compounds present in barley and were found to exhibit the highest binding affinity to both protease and polymeraseDas et al. [136]2020In silicoMolecular dockingFlavonoid-based phytochemicals of Calendula officinalisMain proteaseRutin, isorhamnetin-3-O-β-D, calendoflaside, narcissin, calendulaglycoside B, calenduloside, and calendoflavoside have better binding energy than the native ligandRutin and caledoflaside showed better stability, compactness, and flexibilityDebnath et al. [137]2020In silicoSequential E-pharmacophore and structure-based virtual screening (VS)113687 number of commercially available natural compoundsADP-ribose phosphatase6 potential inhibitors having good binding affinity towards active sitesCommercially availableDev and Kaur [138]2020In silicoMolecular dockingEucalyptus essential oilMain proteaseJensenone may represent potential treatment potential to act as main protease inhibitorDuru et al. [139]2021In silicoMolecular dockingOil of Nigella sativa seedReplicase polyprotein 1a, RNA-binding protein of NSP9, ADP ribose phosphatase of NSP3, 3-chymotrypsin-like protease 3CLpro, and RNA-dependent RNA polymerase RdRp, and ACE2-angiotensin-converting enzyme from the Homo sapiensThe binding affinity of caryophyllene oxide was the highest on NSP9 and RdRp targets, whileα-bergamotene gave the best binding affinity on RPIA target. The binding affinity of β-bisabolene on the ACE2 was almost the same as remdesivirEl-Demerdash et al. [140]2021In silicoMolecular dynamic simulations, molecular docking15 guanidine alkaloidsMain protease (Mpro) (PDB ID: 6lu7), spike glycoprotein (PDB ID: 6VYB), nucleocapsid phosphoprotein (PDB ID: 6VYO), membrane glycoprotein (PDB ID: 6M17), and a nonstructural protein (nsp10) (PDB ID: 6W4H)Crambescidin 786 [5] and crambescidin 826 had the highest binding affinities. The examined 15 alkaloids especially 5 and 13 showed promising docking, ADMET, toxicity, and MD resultsElekofehinti et al. [141]2020In silicoMolecular docking studies, molecular dynamics, and ADME/Tox50,000 natural compounds retrieved from IBS databasePapain-like proteaseSTOCK1N-69160 [(S)-2-((R)-4-((R)-2-amino-3-methylbutanamido)-3-(4-chlorophenyl) butanamido)propanoic acid hydrochloride] has been proposed as a novel inhibitor against COVID-19 PLproEl‐Hawary et al. [142]2021In silicoMolecular docking (a combination of metabolomics and in silico approaches)A. terreus, the endophytic fungus associated with soybean rootsMain proteaseAspergillide B1 and 3α-hydroxy-3,5-dihydromonacolin L were found to be potent anti-COVID-19 drug candidatesEmirik [143]2020In silicoMolecular docking, MM-GBSA-based predictions, and molecular dynamicsTurmeric contentsSARS-CoV-2 vital proteinsTurmeric spice has the potential to inhibit the SARS-CoV-2 vital proteins and can be used a therapeutic or protective agent against SARS-CoV-2 via inhibiting key protein of the SARS-CoV-2. Compounds 4, 23, and 6 are the most prominent inhibitor for the main protease, the spike glycoprotein, and RNA polymerase of virus, respectivelyFakhar et al. [144]2020In silicoStructure-based pharmacophore modeling, virtual screening-based PHASE screen score, molecular modelingAnthocyanin derivativesMain protease6 best anthocyanin-derived natural compounds, which could be used as promising lead compounds against main protease SARS-CoV-2Falade et al. [145]2021In silicoMolecular dockingSaponins and tanninsMain proteaseEllagic acid, arjunic acid, theasapogenol B, and euscaphic acid as potential inhibitors of SARS-CoV-2 (Mpro) with better pharmacokinetics and bioavailability compared with remdesivirFitriani et al. [146]2020In silicoMolecular dockingPhytochemical compounds (Moringa oleifera, Allium cepa, Cocos nucifera, Psidium guajava, and Eucalyptus globulus)Main proteaseOleanolic acid in Allium cepa,α-tocotrienol in Cocos nucifera, asiatic acid in Psidium guajava, and culinoside in Eucalyptus globulus were the most recommended compound in each medicinal plantOleanolic acid in Allium cepa found as a potential inhibitor of COVID-19 MproGangadevi et al. [147]2021In silicoMolecular dynamic simulations, molecular dockingLibrary of natural compoundsHost ACE2 receptor with spike RBD domain of SARS-CoV-2Kobophenol A, identified through docking studies, is the first compound that inhibits SARS-CoV-2 binding to cells through blocking S1-RBD to the host ACE2 receptor and thus may serve as a good lead compound against COVID-19Gangarapu et al. [148]2020In silicoMolecular docking online pkCSM and SwissADME Web serverPhytoconstituents of Siddha official formulation kabasura kudineer and novel herbal preparation—JACOMSpike protein37 compounds were screened, and of these, 9 compounds showed high binding affinity against spike proteinSNACK-V formulations could be used for effective treatment of COVID-19Ghosh et al. [149]2020In silicoMolecular dynamic simulations, molecular docking8 polyphenols from green teaMain protease3 polyphenols (epigallocatechin gallate, epicatechin gallate, and gallocatechin-3-gallate) interact strongly with one or both catalytic residues (His41 and Cys145) of main proteaseGhosh et al. [150]2021In silicoMolecular dynamic simulations, molecular docking, MM-GBSA analysisJusticia adhatoda alkaloidsMain protease1 alkaloid (anisotine) had interaction with both the catalytic residues (His41 and Cys145) of Mpro and exhibited good binding affinity (−7.9 kcal/mol)More potent Mpro inhibitor than the two previously recommended antiviral drugs (lopinavir and darunavir)Gorla et al. [151]2020In silicoMolecular dockingEssential flavonoidsSARS-CoV-2 spike glycoprotein receptor-binding domain (RBD-S) and host angiotensin-converting enzyme-2 protease domain (PD-ACE2)Biochanin A and silymarin bind significantly at the active sites of RBD-Sand PD-ACE2Gurung et al. [152]2020In silicoVirtual screening, molecular dockingAntiviral compounds from plantsMain proteaseBonducellpin D was identified as the best lead molecule, which shows higher binding affinityGyebi et al. [153]2021In silicoMolecular docking, ADME/Tox, and Lipinski filter analysisAfrican plants derived alkaloids and terpenoidsMain protease4 nontoxic, druggable plant-derived alkaloids (10-hydroxyusambarensine and cryptoquindoline) and terpenoids (6-oxoisoiguesterin and 22-hydroxyhopan-3-one)Elwakil et al. [154]2021In silicoGas chromatography/mass spectrometry analysis molecular dockingEgyptian propolisRNA-dependent RNA polymerase, spike protein S1, and main proteaseOctatriacontyl pentafluoropropionate is well oriented inside the enzyme pockets, in addition to an excellent binding manner with the active site of the target macromoleculesMenoufia propolis could be a promising candidate in the combat against the pandemic COVID-19Hasan et al. [155]2020In silicoMolecular dockingCompounds present in the plant Solanum surattenseMain protease13 phytochemicals were studied, eight showed very strong binding affinities to main protease, and four showed moderate to strong binding affinitiesHashem [156]2020In silicoMolecular dockingHoneybee and propolisMain protease6 main compounds possess high binding energy with the receptor active site of the main proteaseIbrahim et al. [157]2020In silicoMolecular dynamic simulations, molecular docking, MM-GBSA analysisMolPort database that contains over 100,000 natural productsMain protease9 potent natural products with binding affinities (ΔG binding) >−48.0 kcal/mol four bis([1, 3]dioxolo)pyran-5-carboxamide derivatives were identified as potential drug candidatesMolPort-004-849-765, MolPort-000-708-794, MolPort-002-513-915 and MolPort-000-702-646 are bis([1,3]dioxolo)pyran-5-carboxamide derivativesIbrahim et al. [158]2020In silicoMolecular dynamic simulations, molecular dockingMetabolites present in several common spicesMain proteaseHigh potency of salvianolic acid A and curcumin as main protease inhibitorsSalvianolic acid A as an in silico natural product inhibitor against the SARS-CoV-2 main proteaseIsa et al. [159]2020In silicoDocking and molecular dynamic (MD) simulationExtracts of Zingiber officinale and Anacardium occidentaleMain proteaseSix compounds had good binding energies. CID_9910474 and CID_10503282 had a better stability when compared to other selected phytochemicalsIstifli et al. [160]2020In silicoMolecular dynamics and molecular mechanic Poisson–Boltzmann surface area (MM/PBSA) methods23 phytochemicals belonging to different flavonoid subgroupsSpike glycoprotein cellular proteases (transmembrane serine protease 2 (TMPRSS2), cathepsin B and L (CatB/L)).(−)-Epicatechin gallate interacted strongly with all the proteins studiedEpicatechin gallate can be evaluated as a candidate molecule in drug development studies against 2019-nCoV since it was not the substrate of P-gp (P-glycoprotein), did not inhibit any of the cytochrome Ps, and did not show AMES toxicity or hepatotoxicity on eukaryotic cellsJan et al. [161]2021In silicoCell-based infection assay molecular modeling2,855 small molecules and 190 traditional herbal medicinesMain protease RNA-dependent RNA polymeraseMefloquine, nelfinavir, and extracts of Ganoderma lucidum (RF3), Perilla frutescens, Mentha haplocalyxJo et al. [162]2020In silicoDockingFlavonoidsMain proteaseBaicalin, herbacetin, and pectolinarin have been discovered to block the proteolytic activity. Baicalin showed an effective inhibitory activity against main proteaseJoshi et al. [163]2021In silicoDocking7100 moleculesMain proteaseSeveral natural molecules such asδ-viniferin, myricitrin, taiwanhomoflavone A, lactucopicrin 15-oxalate, nympholide A, afzelin, biorobin, hesperidin, and phyllaemblicin B strongly binds to main proteaseThese molecules also showed strong binding with other potential targets of SARS-CoV-2 infection such as viral receptor human angiotensin-converting enzyme 2 (hACE2) and RNA-dependent RNA polymerase (RdRp)Junior et al. [164]2021In silicoDocking and molecular dynamic (MD) simulationLapachol(1,4-naphthoquinone)SARS-CoV-2 nonstructural proteins (nsps)Lapachol derivatives VI and IX demonstrated the strongest bindingLapachol derivatives are potential ligands for SARS-CoV-2 Nsp9Kar et al. [165]2020In silicoMolecular docking molecular dynamic simulations and analysis of MM-PBSA energyIndian plants including Justicia adhatoda, Ocimum sanctum, and Swertia chirataSpike protein, main protease enzyme Mpro, and RNA-dependent RNA polymerase (RdRp)Anisotine against SARS-CoV-2 spike and Mpro proteins and amarogentin against RdRp as potential inhibitorsKhalifa et al. [166]2020In silicoMolecular docking modeling structural-relationship activity10 anthocyaninsMain proteasePhacelianin, gentiodelphin, cyanodelphin, tecophilinLeading molecules for further optimization and drug development process to combat COVID-19Khalifa et al. [167]2020In silicoMolecular operating environment molecular docking19 structural different hydrolysable tanninsMain proteasePedunculagin, tercatain, and castalinKhan et al. [168]2020In silicoMolecular dockingMarine natural productsMain proteaseC-1 (PubChem CID 11170714) exhibited good activityKiran Raj et al. [169]2020In silicoMolecular dockingC-Phycocyanin of Spirulina platensisNonstructural proteins 12C-Phycocyanin inhibits the active site of nsp12Krupanidhi et al. [170]2020In silicoMolecular docking molecular dynamic simulation ADME along with toxicity analysisPhytochemical constituents of Tinospora cordifoliaMain proteaseTinospononeKumar et al. [171]2020In silicoMolecular docking molecular dynamic simulations and analysis of MM-PBSA energyNovel natural metabolitesMain proteaseUrsolic acid, carvacrol, and oleanolic acid couldKumar et al. [172]2021In silicoMolecular docking, ADMET, and molecular dynamic simulationsPhytoconstituents from natural herbsMain proteaseLaurolitsineLaurolitsine, an active constituent of roots of Lindera aggregataKumar et al. [173]2021In silicoMolecular docking molecular dynamic simulations and analysis of MM-PBSA energyStrychnos nux-vomicaMain proteaseDemethoxyguiaflavine, strychnoflavineLi et al. [174]2021In silicoNetwork pharmacology and in vitro experiment verification molecular dockingMaxing Shigan decoction (MXSGD)ACE2, Mpro, and RdRpThe components with strong potential affinity (top 10) with ACE2, Mpro, and RdRp are mainly from Glycyrrhiza uralensis (Chinese name: Gancao) and Semen armeniacae amarum (Chinese name: Kuxingren). Among them, amygdalin was selected as the optimal candidate component binds to all three key targets, and euchrenone, glycyrrhizin, and glycyrol also exhibited superior affinity interactions with ACE2, Mpro, and RdRp, respectivelyMulticomponent, multitarget, and multi-approach interventionMaiti and Banerjee [175]2021In silicoBioinformatic molecular dockingTea flavonoid catechin productsSpike glycoproteinsEpigallocatechin gallate and theaflavin gallate interact better than hydroxychloroquineMahmud et al. [176]2021In silicoMolecular docking molecular dynamic simulation MM-GBSA scores3063 compounds from more than 200 plants from the Asian regionMain proteaseCurcumin, gartanin, robinetinMahmud et al. [177]2020In silicoMolecular docking molecular dynamic simulation MM-GBSA scoresPlant-derived natural compoundsMain proteaseCarinol, albanin, myricetinMesli et al. [178]2021In silicoMolecular docking molecular dynamic simulationsLeaves of Corchorus olitorius Linn.Angiotensin-converting enzyme 2Méthyl-1,4,5-tri-O-caféoyl quinate has a stronger bond, high affinity, and gives the best docking scores compared with the co-crystallized inhibitor (PRD_002214) of the enzyme ACE2Mohammadi et al. [179]2020In silicoMolecular dockingMarine polypeptides were isolated from the hydrolysate of Pacific oysterMain proteaseThe score of Leu-Leu-Glu-Tyr-Ser-Ileu ligand was higher than remdesivirPacific oyster compounds may have the potency to be evolved as an anti-COVID-19 main proteaseMurugan et al. [180]2020In silicoMolecular docking molecular dynamic simulation MM-GBSA scoresAndrographis paniculata phytochemicals3 nonstructural proteins (3 L main protease (3CLpro), papain-like proteinase (PLpro) and RNA-directed RNA polymerase (RdRp)), and a structural protein (spike protein (S))Neoandrographolide (AGP3) has shown promising binding affinity towards all the four targetsNaik et al. [181]2020In silicoMolecular docking molecular dynamic simulation ADME propertiesNatural product activity and species source (NPASS) databaseEndoribonuclease exoribonuclease RNA-dependent RNA polymerase (RdRp) methyltransferase and main protease21 compounds showed maximum docking scores NPC214620, NPC52382, and NPC270578 are targeting five, four, and three-drug targets, respectivelyMultitarget-based drug designNarkhede et al. [182]2020In silicoMolecular docking molecular dynamic simulationsNatural productsMain proteaseGlycyrrhizin, bicyclogermacrene, tryptanthrin,β-sitosterol, indirubin, indican, indigo, hesperetin, chrysophanic acid, rhein, berberine, and β-caryophylleneInteractions with the COVID-19 main protease were highest in the case of glycyrrhizin and rheinNivetha et al. [183]2020In silicoMolecular docking molecular dynamic simulation MM-PBSASeselin purified from the leaf extracts of Aegle marmelosSpike protein S2, main protease, and free enzyme of the SARS-CoV-2Seselin had inhibitory potential over multiple SARS-CoV-2 targetsOgunyemi et al. [184]2020In silicoMolecular docking molecular dynamic simulation ADME properties226 bioactive compounds from African herbs and medicinal plantsRNA-dependent RNA polymeraseDrugable alkaloids (10′-hydroxyusambarensine, cryptospirolepine, strychnopentamine) and flavonoids (usararotenoid A and 12α-epi-millettosin)Padhi et al. [185]2021In silicoMolecular docking ADME properties415 natural metabolites isolated from several plants, fungi, and bacteriaSpike glycoprotein (S1) and the main proteaseFusaric acid, jasmonic acid, jasmonic acid methyl ester, putaminoxin, putaminoxins B and D, and stagonolide K were predicted to have considerable (ADME) and safety indicesJasmonic acid and putaminoxins B and D bind best to main proteasePandey et al. [186]2020In silicoMolecular docking molecular dynamic simulation ADME properties10 potential naturally occurring compounds (flavonoids/non-flavonoids)Spike glycoproteinFisetin, quercetin, and kaempferol consist of drug-likeness propertyKumar et al. [187]2020In silicoMolecular dockingKabasura kudineer and thonthasura kudineer are two Siddha formulationsSpike glycoproteinCucurbitacin B (−112.09), cardiofolioside (−111.5), apigenin (−98.84), and pyrethrin (−92.98) were observed as more effective with less bind energiesKabasura kudineer could be a potential Siddha medicine for COVID-19Rahman et al. [188]2021In silicoMolecular docking ADMET propertiesRutinMain protease (Mpro), RNA-dependent RNA polymerase (RdRp), papain-like protease (PLpro), and spike (S) proteinSignificant binding of rutin with Mpro, RdRp, PLpro, and S proteins. Main protease exhibited the strongest binding affinityOptimal solubility, nontoxic, and noncarcinogenic propertiesRahman et al. [189]2020In silicoMolecular operating environment (MOE) ligand-based pharmacophore approach and a molecular docking-based screeningNatural product activity and species source (NPASS)Type II transmembrane serine protease (TMPRSS2)85 compounds with molecular docking comparable to or greater than that of the standard inhibitor (camostat mesylate) were identified. The top 12 compounds with the most favorable structural features were studied. The low-molecular-weight compound NPC306344 showed significant interaction with the active site residues of TMPRSS2Rakib et al. [190]2020In silicoMolecular dockingBioactive phytocompounds isolated from Tinospora crispaMain proteaseThe top nine hits might serve as potential anti-SARS-CoV-2 lead molecules, with three of them exerting biological activityRamadhan et al. [191]2020In silicoMolecular dockingEtlingera elatior plantMain proteaseErgosterol peroxide sitostenoneRangsinth et al. [192]2021In silicoMolecular docking ADMET propertiesNatural products isolated from edible and medicinal mushroomsMain protease6 of 25 compounds are the best drug-like property candidates, including colossolactone VIII, colossolactone E, colossolactone G, ergosterol, heliantriol F, and velutinRivero-Segura et al. [193]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesMexican natural productsAgainst the SARS-CoV-2Quercetin, riolozatrione, and cichoriin target the key proteins of SARS-CoV-2Cichoriin reaches higher lung levels (100 mg/kg, IV); therefore, it may be considered in developing therapeutic toolsSelvaraj et al. [194]2020In silicoHomology modeling and molecular dynamic (MD) simulation MM/GBSA, MD simulations, and PCA calculationsTraditional Chinese medicine (TCM) databaseNsp 14 guanine-N7 methyltransferase (N7-MTase)TCM 57025, TCM 3495, TCM 5376, TCM 20111, and TCM 31007 are the compounds from the TCM database, which can occupy and interact nicely with the substrate-binding site of N7-MTaseSharma and Kaur [195]2021In silicoMolecular docking, protein interaction calculator ADME studies12 bioactive molecules present in essential oils of eucalyptus plant leavesSpike (S) proteinToruatoneSharma [196]2020In silicoMolecular docking, protein interaction calculatorEucalyptol (1,8 cineole), an essential oil component from eucalyptus oilMain proteaseEucalyptol may represent potential treatment potential to act as main protease inhibitorEffective binding of eucalyptol to COVID-19 proteinaseSharma and Kaur [197]2020In silicoMolecular docking, protein interaction calculatorJensenone, an essential oil component from eucalyptus oilMain proteaseJensenone may represent potential treatment potential to act as main protease inhibitorShawan et al. [198]2021In silicoPharmacophore study molecular docking molecular dynamic simulation ADME properties43 flavonoids of 7 different classesAgainst the SARS-CoV-2Luteolin and abyssinone II were found to develop successfully docked complex within the binding sites of target proteinsSindhu et al. [199]2020In silicoMolecular dockingClerodendrum paniculatum leavesMain proteaseClerodolSingh et al. [200]2021In silicoMolecular docking and structural dynamic studiesTea (Camellia sinensis) polyphenolsNonstructural protein 16 (NSP16)Theaflavin compound demonstrated lower binding free energy in comparison with the standard molecule sinefunginSingh et al. [201]2020In silicoMolecular docking and structural dynamic studies molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) ADME propertiesPolyphenolsRNA‐dependent RNA polymerase (RdRp)EGCG, theaflavin (TF1), theaflavin-3′-O-gallate (TF2a), theaflavin-3′-gallate (TF2b), theaflavin 3,3′-digallate (TF3), hesperidin, quercetagetin, and myricetin strongly bind to the active site of RdRpEGCG, TF2a, TF2b, and TF3 can inhibit RdRp and represent an effective therapy for COVID-19Srimathi et al. [202]2020In silicoMolecular dockingTraditional herbal medicine: apo-quinine, catechin, cinchonidine, cinchonine, cupreidine, epicatechin, epiprocurcumenol, epiquinine, procurcumenol, quinidine, quinine, zedoaronediol, procurcumadiolAgainst the SARS-CoV-2Epicatechin, apo-quineSubbaiyan et al. [203]2020In silicoMolecular dockingActive constituents present in common herbsSpike (S) proteinEpigallocatechin gallate (EGCG) was found to have the highest binding affinity with the viral S protein, followed by compounds, “F” (curcumin), “D” (apigenin), and “E” (chrysophanol)Green teaSurti et al. [204]2020In silicoMolecular docking molecular dynamic simulationsIlimaquinone (marine sponge metabolite)Spike receptor-binding domain, RNA-dependent RNA polymerase, Nsp10, Nsp13, Nsp14, Nsp15, Nsp16, main protease, and papain-like proteaseIlimaquinone exhibited promising inhibitory potential against all the SARS-CoV-2 target proteins, as evident from the binding energiesMost promising inhibitory candidate against the SARS-CoV-2 papain-like proteaseTao et al. [205]2020In silicoNetwork pharmacology and molecular docking.Huashi Baidu formula (HSBDF): ChineseAgainst the SARS-CoV-2Baicalein and quercetin were the top two compounds of HSBDF, which had high affinity with ACE2Regulate multiple signaling pathways through ACE2Umar et al. [206]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesAzadirachta indica, Mangifera indica, and Moringa oleifera: African plantsMain proteaseMost of the active phytocomponents of the study plants exhibited relative inhibitory potentials against main protease and preferred pharmacological features when compared with hydroxychloroquineCaffeic acid, chlorogenic acid, catechin, ellagic acid, gallic acid, etc.Umesh et al. [207]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesChemical compounds from Indian spicesMain proteaseCarnosol exhibited the highest binding affinity for arjunglucoside-I and rosmanol showed a strong and stable binding affinity with favorable ADME propertiesYang et al. [208]In silico2020High-throughput virtual screeningNatural Products Research Laboratories (NPRL)Main proteaseCurcuminoid derivatives (including NPRL334, NPRL339, NPRL342, NPRL346, NPRL407, NPRL415, NPRL420, NPRL472, and NPRL473) display strong binding affinity to COVID-19 3Lpro polyproteinNPRL-334 revealed the strongest binding affinityYu et al. [209]2020In silicoMetascape analysis protein docking molecular dockingMongolian medicineSARS-CoV-2 S protein RBD domain253 active components were predicted. Phillyrin and chlorogenic acid can effectively prevent the combination of SARS-CoV-2 S protein and ACE2 at the molecular levelZhang et al. [210]2020In silicoMolecular docking molecular dynamic simulation ADME property network pharmacology analysisChinese herbal medicinesAnti-2019-nCoV activity13 compounds that exist in traditional Chinese medicines were found to have potential anti-2019-nCoV activity. 125 Chinese herbs were found to contain 2 or more of these 13 compounds. Of these 125 herbs, 26 are classically cataloged as treating viral respiratory infectionsRegulating viral infection, immune/inflammation reactions, and hypoxia responseTable 3
Summary of other studies with in silico method.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAbu-Melha et al. [211]2020In silicoMolecular docking combined with molecular dynamic simulation (MDS)Hydrazones, pyrazoles, and pyrazines bearing thiazole moietyMain proteaseThe average binding affinities of the compounds 3a, 3b, and 3c (−8.1 ± 0.33 kcal/mol, −8.0 ± 0.35 kcal/mol, and −8.2 ± 0.21 kcal/mol, respectively) are better than that of the positive control nelfinavirAghaee et al. [212]2021In silicoPharmacophore model molecular docking combined with molecular dynamic simulation (MDS), MM/PBSA, ADME studiesPharmit websiteMain proteaseML188, nelfinavir, lopinavir, ritonavir, andα-ketoamideAhmad et al. [213]2021In silicoStructure-based virtual screening (SBVS) of ASINEX antiviral library, molecular dynamic (MD) simulationsASINEX antiviral libraryMain proteaseSCHEMBL12616233, SCHEMBL18616095, and SCHEMBL20148701 compounds conformation with main protease show good stability after initial within active cavity moves, a rich intermolecular network of chemical interactions, and reliable relative and absolute binding free energiesBBB_26580140 lead and its similar analogs to be explored in vivo lead moleculesAhmed et al. [214]2020In silicoMolecular docking, molecular dynamics, and structure-activity relationship76 prescription antiviral drugsRNA-dependent RNA polymerase (RdRp) and main protease (Mpro)Raltegravir, simeprevir, cobicistat, and daclatasvir have higher binding energy and strong interaction with active sites of the receptor proteins (with a precision of 85%)Alabboud and Javadmanesh [215]2020In silicoMolecular docking combined with molecular dynamic simulation88 conventional drugs, 16 vitamins, and 63 natural (plant)Main proteaseVarious vitamins (B9, A, K, and E vitamins) exhibited a significantly strong interaction with the studied receptor. Pleconaril, adefovir dipivoxil, and stavudine in addition to plant-based compounds such as curcumin (Curcuma longa), anolignan A (Anogeissus acuminata), and phyllamyricin B (Phyllanthus myrtifolius) had strong ligand-protein interactionsAlamri et al. [216]2020In silicoStructure-based virtual screening coupled with all-atom molecular dynamic (MD) simulationsProtease inhibitors database composed of ∼7,000 compoundsPapain-like proteaseADM_13083841, LMG_15521745, and SYN_15517940 showed stable conformation and interacted well with the active residues of papain-like proteaseAlexpandi et al. [217]2020In silicoMolecular docking113 quinoline drugsMain protease, RNA-dependent RNA polymerase (RdRp) inhibitors spike-RBD-ACE2 inhibitorElvitegravir and oxolinic acid are able to interact with the NTP entry channel and thus interfere with the RNA-directed 5′–3′ polymerase activity of SARS-CoV-2 RdRp. Rilapladib is the only quinoline that can interrupt the spike-RBD-ACE2 complexQuinoline, 1,2,3,4-tetrahydro-1-[(2-phenylcyclopropyl)sulfonyl]-trans-(8CI), saquinavir, elvitegravir, oxolinic acid, and rilapladib are suggested for the treatment of COVID-19Al‐Sehemi et al. [218]2020In silicoMolecular docking, MD simulation, and molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) resultsPhenyl furoxan, an exogenous nitric oxide donorMain proteaseSpiro-isoquinolino-piperidine-furoxan moieties can be used as an effective ligand for main protease inhibition due to the presence of key isoquinolino-piperidine skeleton with additional NO effectAl-Shar’i [219]2020In silicoMolecular docking, MD simulation, and molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) resultsDifferent databasesMain protease9 compounds with different chemotypesBadavath et al. [220]2020In silico/in vitroMolecular docking, molecular dynamics, and structure-activity relationship studiesScreening of 118 compounds with 16 distinct heterocyclic moieties in comparison with 5 natural products and 7 repurposed drugsMain proteaseOxidiazoles (A2 and A4) derivatives have the best docking scores. Structure-activity relationship studies showed a good comparison with a known active main protease and repurposed drug ebselen with an IC50 value of −0.67 μMBarros et al. [221]2020In silicoMolecular docking24 ligandsSARS-CoV-2 receptors, Nsp9 replicase, main protease (Mpro), NSP15 endoribonuclease, and spike protein (S protein) interacting with human ACE2Antimalarial drug metaquine and anti-HIV antiretroviral saquinavir interacted with all the studied receptorsBasit et al. [222]2020In silicoProtein-protein docking and molecular dynamic simulationTruncated version of human ACE2 (tACE2)S GlycoproteintACE2 provides a high-affinity protein inhibitor for S glycoproteinBattisti et al. [223]2020In silicoPharmacophore‐based screening, docking consensus approach (DCA), molecular dynamic simulations, common hit approach (CHA)Aldrich Market Select (AMS) database from ChemNavigator/Sigma‐Aldrich with over 8 million unique chemical structuresAgainst SARS-CoV-210 compounds with high coronavirus inhibition potentialLead moleculesBenítez-Cardoza and Vique-Sánchez [224]2020In silicoMolecular docking500,000 compoundsPotential inhibitors of the interaction between ACE2 and SARS-CoV-2 (RBD)20 compounds were determined by docking focused on the region of interaction between ACE2 and RBDCava et al. [225]2020In silicoGene Ontology and enrichment analysis protein-protein interaction (PPI) network virtual screening method—Against SARS-CoV-2A protein-protein interaction network of 193 genes, 22 interactions, and 36 potential drugs for future treatment strategies including nimesulide, fluticasone propionate, thiabendazole, photofrin, and didanosineOnly didanosine is a real antiviral drug, while the others are mostly anti-inflammatoryChen et al. [226]2020In silicoCrystal structure, virtual screening7173 purchasable drugs (drugs-lib), with 4574 unique compounds and their stereoisomersMain protease16 candidates for consideration, ledipasvir velpatasvirChoudhary et al. [227]2020In silicoMolecular docking, MM-GBSA predictive binding energy calculations, and molecular dynamic simulation15,754 natural and synthetic compoundsMain proteaseCompound 2 (molecular bank code AAA396) and compound 3 (molecular bank code AAD146)Chunduru et al. [228]2021In silicoMolecular dockingNovel drug-like inhibitors for COVID-19Main proteaseStructure 61 was found to be more stable and can be further assessed for their antiviral activity to combat COVID-19Coelho et al. [229]2020In silicoBiochemical high-throughput screeningCompound library containing known drugs, bioactive molecules, and natural productsMain proteaseOrganomercuric compounds thimerosal and phenylmercuric acetate, benzophenone, Evans blue, a sulfonic acid-containing dyeDai et al. [230]2020In silicoStructure-based design—Main proteaseDesigned and synthesized two lead compounds (11a and 11b) targeting main protease. Both exhibited excellent inhibitory activityde lima Menezes and da Silva [231]2020In silicoMolecular dynamic simulations, molecular dockingDrugBank databaseNonstructural protein 1 (nsp1)Tirilazad, phthalocyanine, and Zk-806450 showed better energy score than control molecules that have in vitro activity against nsp1 from SARS-CoV-2Tirilazad, phthalocyanine, and Zk-806450Debnath et al. [232]2020In silicoPharmacophores studies, structure-based virtual screening, molecular dynamic (MD) simulationDrug molecule information retrieved from DrugBankMain proteaseDB07456 and DB13592 displayed a similar type of binding interaction with co-ligands and remdesivir, and the predicted Ki values of 2 inhibitors were found to be superior to remdesivirDi Micco et al. [233]2021In silicoMolecular docking, MM-GBSA-based predictions, and molecular dynamicsZonulin inhibitor larazotide acetate (also called AT1001)Main proteaseAT1001, besides its well-demonstrated effect in ameliorating mucosal permeability in ALI/ARDSEl Hassab et al. [234]2021In silicoStructure-based virtual screening, molecular dynamic simulation, and MM-PBSA approaches48 million drug-like compounds of the ZINC databaseSARS-CoV-2 2′-O-methyltransferase (nsp16)Compound 11 as the best potential nsp16 inhibitor herein identified, as it displayed a better stability and average binding free energy for the ligand-enzyme complex compared to sinefunginElmessaoudi-Idrissi et al. [235]2020In silicoIn silico screening, molecular docking, and dynamic approaches5000 compounds of the ZINC databaseMain proteaseThe prominent drug-like and potent inhibitory compounds are 2-[2-(2-aminoacetyl) aminoacetyl] amino-3-(4-hydroxyphenyl)-propanamide (ZINC000004762511), 6′-fluoroaristeromycin (ZINC000001483267), and cyclo(L-histidyl-L-histidyl) (ZINC000005116916) scaffoldsFeitosa et al. [236]2020In silicoMolecular docking—Main proteaseMelatonin can have response potential in early stages for its possible effects on ACE2 and main protease, although it is also promising in more severe stages of the disease for its action against hyper-inflammationDo not confirm antiviral activity, but can rather be used as a basis for further preclinical and clinical trialsGurung et al. [237]2021In silicoVirtual screening, molecular docking1,36,191 moleculesSpike (S) protein receptor-binding domain (RBD) to the host cell surface receptor, angiotensin-converting enzyme 2 (ACE2)ZINC33039472 exhibited binding free energy value lower as compared to the control (emodin) with a higher contribution by gas-phase energy and van der Waals energy to the total binding free energyHaider et al. [238]2020In silicoComputer-aided drug design (CADD) molecular dockingZINCPharmerMain protease∼200 compounds were identified as having strong interaction with Mpro (ZINC20291569, ZINC90403206, ZINC95480156) that showed the highest binding energyHall and Ji [239]2020In silicoHomology modeling molecular dockingZINC15 database: 3447 entriesSpike glycoprotein and main proteaseZanamivir, indinavir, saquinavir, and remdesivir are among the exciting hits of main proteinaseFlavin adenine dinucleotide (FAD) adeflavin, B2 deficiency medicine, and coenzyme A, a coenzyme may also be potentially used for the treatment of SARS-CoV-2 infectionsHavranek and Islam [240]2020In silicoDocking and molecular dynamics2692 protease inhibitor compoundsMain proteasePhenyltriazolinones (PubChem ID: 104161460) and allosteric HCV NS5B polymerase thumb pocket 2 (PubChem ID: 163632044) have shown antiviral activity and also have high affinity towards the main proteaseIbrahim et al. [241]2021In silicoMolecular dynamic simulations, molecular docking, MM-GBSA18 anti-COVID-19 drug candidates against SARS-CoV-2 main proteaseMain proteasePromising binding affinities of TMC-310911 and ritonavirJaiswal and Kumar [242]2020In silicoDocking studies and molecular dynamic simulationDesigned a protein inhibitorSpike (S) glycoproteinThe proposed inhibitor ΔABP-D25YJamalan et al. [243]2021In silicoDocking and molecular dynamic (MD) simulationVirtual screening based on GRL-0617Papain-like proteinase5-(aminomethyl)-2-methyl-N-[(1R)-1-naphthalen-1-ylethyl]benzamide outperformed GRL-0617 in terms of binding affinity (−9.7 kcal/mol). 2-(4-fluorobenzyl)-5-nitro-1H-isoindole-1,3(2H)-dione, 3-nitro-N-[(1r)-1-phenylethyl]-5-(trifluoromethyl)benzamide, 5-acetamido-2-methyl-N-[(1S)-1-naphthalen-1-ylethyl]benzamideJin et al. [244]2020In silicoStructure-assisted drug design, virtual drug screening, and high-throughput screening10,000 compoundsMain proteaseEbselen, disulfiram, tideglusib, carmofur, shikonin, PX-12Ebselen also exhibited promising antiviral activity in cell-based assaysKanhed et al. [245]2021In silicoSystematic virtual screening approachASINEX BioDesign library approved drug libraryMain proteaseRitonavir, nelfinavir, and saquinavir were predicted to be the most potent Mpro inhibitors. 20 molecules (pyrazoles, cyclic amides, pyrrolidine-based compounds, and miscellaneous derivatives)Kavitha et al. [246]2020In silicoMolecular docking molecular dynamic simulations1000 protease-inhibitor-like compounds available in the ZINC databaseMain protease1,2,4 triazolo[1,5-a] pyrimidin-7-onesKrishnan et al. [247]2020In silicoMolecular docking3978 compounds with potential antiviral activityEndoribonuclease (NSP15)8 compounds with good docking score and docking energy e.g., Z595015370, Z1343129850, and Z2760938911Kumar et al. [248]2021In silicoMolecular docking molecular dynamic simulation molecular mechanic Poisson–Boltzmann surface area approachesMillion molecules and natural compound databasesMain proteaseThree compounds namely ZINC14732869, ZINC19774413, and ZINC19774479 displayed better binding affinitiesKumar et al. [249]2020In silicoMolecular docking molecular dynamic simulations13 approved antiviral drugsMain proteaseIndinavir was described as a lead drug. Indinavir possesses an important pharmacophoreNovel compound 16(hydroxyethylamine derivative) suitability as a strong candidate for therapeutic discovery against COVID-19Kwarteng et al. [250]2020In silicoBioinformatic approach molecular docking molecular dynamic simulations—Nucleocapsid (N) proteinZidovudine triphosphate, an anti-HIV agent, as a potential inhibitor of the N-terminal domain of SARS-CoV2 N proteinLi et al. [251]2021In silicoMolecular docking21 antiviral, antifungal, and anticancer compoundsPapain-like proteaseNeobavaisoflavoneMathpal et al. [252]2020In silicoMolecular docking molecular dynamic simulation MM-PBSA3180 FDA-approved drugs from “the ZINC database”Main proteaseZINC03831201, ZINC08101052, ZINC01482077, and ZINC03830817Maurya et al. [253]2020In silicoMolecular dockingAntiviral, anti-infectious, and anti-protease compoundsNSP10/NSP16 methyltransferase and main proteaseCyclocytidine hydrochloride, trifluridine adonitol, and meropenem penciclovir bound with a good docking score NSP10/NSP16 methyltransferase complexed with telbivudine, oxytetracycline dihydrate, methyl gallate, 2-deoxyglucose, and daphnetinMohamed et al. [254]2020In silicoMolecular docking12 histone deacetylases (HDACs)Main proteaseRomidepsin and its active form (RedFK)Monajemi and Zain [255]2021In silicoONIOM (own N-layered integrated molecular orbital and molecular mechanics; QM/MM) approachN3, ebselen, disulfiram, tideglusib, carmofur, shikonin, and PX-12Main proteaseN3, ebselen, and PX-12 inhibitorsBetter inhibition from PX-12 than ebselenMotiwale et al. [256]2020In silicoMolecular docking molecular dynamic simulationsPreviously reported SARS-3CL protease inhibitorsMain proteaseN-substituted isatin derivatives and pyrazoloneMutlu et al. [257]2020In silicoStructure-based approach molecular docking molecular dynamic simulationsFDA-approved and investigational drugsNsp12/Nsp8Two drugs, RX-3117 (fluorocyclopentenyl cytosine) and nebivololNaidoo et al. [258]2020In silicoMolecular docking molecular dynamic simulation MM-PBSACyanobacterial metabolitesMain protease (Mpro) and the papain-like protease (PLpro)Deoxycylin, drospermopsinCylindrospermopsin, deoxycylindrospermopsin, carrageenan, cryptophycin 52, eucapsitrione, tjipanazole, tolyporphin, and apratoxin A exhibited promising inhibitory potential against the SARS-CoV-2 Mpro. The compounds cryptophycin 1, cryptophycin 52, and deoxycylindrospermopsin were observed to display encouraging binding energy scores with the PLpro of SARS-CoV-2Olubiy et al. [259]2020In silicoMolecular docking molecular dynamic simulationsApproved drugs, investigational drugs, natural products, and organic compoundsMain proteaseSeveral tyrosine kinase inhibitors, which include a bioflavonoid and steroid hormones, bind best to main proteaseNilotinib, enasidenib, afatinib, ertapenem, phthalocyanine, hypericin, amrubicin, theacitrin A, theaflavin, amentoflavone, epigallocatechin gallate, glabrolide, cortisol, estradiol, testosteroneÖzdemir et al. [260]2020In silicoMolecular docking density functional theory (DFT) ADME-Tox42 coumarin derivativesMain protease6,7-Dihydroxy-3-phenylcoumarin derivatives gave relatively higher scores, and for all coumarins, and 4-trifluoromethylphenyl substituted coumarin had the highest scoreOzdemir et al. [261]2020In silicoMolecular docking molecular mechanics Poisson–Boltzmann surface area (MM-PBSA)Coumarin derivativesSpike S1 subunit, NSP5, NSP12, NSP15, and NSP16The highest score (−10.01 kcal/mol) in the coumarin group is 2-morpholinoethan-1-amine substituted coumarinPatel et al. [262]2021In silicoPharmacophore studiesDrug library (having drugs and diagnostic agents, which are approved by FDA or other world authorities) and the ASINEX BioDesign libraryMain proteaseRitonavir, nelfinavir, and saquinavir were predicted to be the most potent Mpro inhibitors 20 molecules categorized into four classes viz. disubstituted pyrazoles, cyclic amides, pyrrolidine-based compounds, and miscellaneous derivativesPeng et al. [263]2020In silicoDrug repositioning through virus-drug association prediction—Against SARS-CoV-2Ribavirin was predicted to be the best small molecular drug, with a higher molecular binding energy with human angiotensin-converting enzyme 2 (ACE2), followed by remdesivir, mycophenolic acid, and chloroquine (−6.29 kcal/mol)Pratama et al. [264]2020In silicoMolecular dockingNovel 5-O-benzoylpinostrobin derivativesMain proteaseThree 5-O-benzoylpinostrobin derivatives each with fluoro, tertiary butyl, and trifluoromethyl substituents at 4-position of benzoyl group showed the lowest free energy of binding value and the highest similarity of ligand-receptor interactions with co-crystallized ligandsPundir et al. [265]2020In silicoPharmacophore-based virtual screening molecular mechanics Poisson–Boltzmann surface area (MM-PBSA)PubChem databaseMain proteaseTwo compounds: PubChem3408741 and PubChem4167619 had the binding free energy of −94.02 kJ·mol−1 and −122.75 kJ·mol−1, respectively, as compared to reference X77 (−76.48 kJ·mol−1)Lead molecules for targeting Mpro enzymeQuimque et al. [266]2020In silicoMolecular docking molecular dynamic simulation ADME properties97 antiviral secondary metabolites from fungiPapain-like protease, RNA-dependent RNA polymerase, main protease, spike glycoprotein, nonstructural protein 15 (nsp15)Two fumiquinazoline alkaloids quinadoline B (19), scedapin C (15), and polyketide isochaetochromin D1 (8)Quinadoline B [19] was predicted to confer favorable ADMET values, high gastrointestinal absorptive probability, and poor blood-brain barrier crossing capacitiesRakib et al. [267]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesSelenium-containing heterocyclic compoundsMain proteaseSelection of the 16 most effective selenocompounds as potential anti-COVID-19 drug candidates. Ethaselen showed potential binding affinitiesRane et al. [268]2020In silicoMolecular docking molecular dynamic simulationsDiarylpyrimidine analogsSpike glycoproteinAP-NP (2-(2-amino-5-(naphthalen-2-yl)pyrimidin-4-yl)phenol), AP-3-OMe-Ph (2-(2-amino-5-(3-methoxyphenyl)pyrimidin-4-yl)phenol), and AP-4-Me-Ph (2-(2-amino-5-(p-tolyl) pyrimidin-4-yl)phenol) from a group of diarylpyrimidine derivatives, which appears to bind at the interface of the hACE2-S complex with low binding free energyRao et al. [269]2020In silicoMolecular docking molecular dynamic simulationsVarious small moleculesMain proteasePyranonigrin A, a secondary fungal metaboliteSalman et al. [270]2020In silicoMolecular docking molecular dynamic simulation ADME propertiesLibrary of immunomodulatory medicinal compounds with antiviral capabilitySARS proteases, spike protein, and nonstructural proteins (NSP-9, 15)6 compounds: arzanol, ferulic acid, genistein, resveratrol, rosmanol, and thymohydroquinoneGood pharmacokinetic properties and low acute toxicity of these compoundsSarma et al. [271]2020In silicoMolecular docking MM-GBSA binding free energy molecular dynamic simulations56,079 compounds from ASINEX and Maybridge libraryRNA-binding N-terminal domain (NTD) of the N proteinZINC00003118440 is a theophylline derivative. Pyrimidone derivatives as possible inhibitors of RNA binding to the N-terminal domain of N protein of coronavirusLead moleculesSepay et al. [272]2020In silicoMolecular docking, bioinformatics, and molecular electrostatic potential ADME studiesBenzylidenechromanones, naturally occurring oxygen heterocyclic compoundsMain protease(Z)-3-(4 ′-chlorobenzylidene)-thiochroman-4-Effective pharmacological propertiesShehroz et al. [273]2020In silicoPharmacophore modeling molecular dockingDrugBank, ZINC, and TIMBLE databasesSpike (S) proteinOnly eight molecules fit the criteria for safe oral drugsLead moleculesSingh and Das [274]2021In silicoMolecular dockingChloroquine (CQ) hydroxychloroquine (HCQ) azithromycinSpike (S) protein main protease host cathepsin L (CTSL) receptor-binding domain (RBD)Azithromycin affinity scores (ΔG) with strong interactions with ACE2, CTSL, Mpro, and RBD. CQ firm bond score with Mpro HCQ and two results (ACE2 and Mpro) were firmly bound to the receptorsStefaniu et al. [275]2020In silicoMolecular docking density functional theory (DFT) computations, drug-likeness assessmentDerivatives of benzoic acidMain protease2,5-dihydroxybenzoic acid (gentisic acid) and octylA combination of the two compounds can be consideredTachoua et al. [276]2020In silicoMolecular docking and structural dynamic study molecular mechanic Poisson–Boltzmann surface area (MM-PBSA) ADMET analysisChloroquine, quinine, nitazoxanide, doxycycline, lymecycline, cetirizine, mizolastine, indinavirMain proteaseLymecycline mizolastineUniyal et al. [277]2020In silicoStructure-based virtual screening molecular docking MM-GBSA binding free energy molecular dynamic simulationsCommercially available chemical librariesMain proteaseCompound AG-690/11203374_1 and AG-690/11203374_2 emerged as the best in silico hits based on the docking, MM-GBSA, dynamics, and ADMET studiesLead moleculesWelker et al. [278]2020In silico/in vitroStructure-activity relationships molecular docking fluorescence-based enzyme‐activity assay Vero E6 cellsA series of rationally designed competitive, noncovalent, nonpeptidic active site‐directed SARS‐CoV PLpro inhibitorsPapain-like cysteine proteases (PLpro)R)‐5‐amino‐2‐methyl‐N‐(1‐(naphthalen‐1‐yl)ethyl) benzamide (2b), which is known to bind into the S3 and S4 pockets of the SARS‐CoV PLpro. Isoindoline as a new class of potent PLpro inhibitorsIC50 value of 2.9 ± 0.2 μMWen et al. [279]2021In silicoStructure-based screening8,820 compoundsMain proteaseTrichostatin AA histone deacetylase inhibitor and an antifungal compoundWhite et al. [280]2020In silicoHomology modeling and molecular dynamics approach molecular docking∼970,000 chemical compoundsHelicase (Nsp13)Nilotinib and lumacaftor have significant activity in inhibiting purified recombinant SARS-CoV-2 helicaseWu et al. [281]2020In silicoMolecular docking11 HIV-1 protease inhibitors, 12 nucleotide-analog inhibitors, 728 approved drugsMain protease RNA-dependent RNA polymeraseRemdesivir shows the best binding energy on RdRp and saquinavir is the best inhibitor of main proteaseZaher et al. [282]2020In silicoDesign, synthesis SAR study molecular dockingNewly synthesized sixteen halogenated triazole compoundsHelicase (Nsp13)The most potent compounds were 4-(cyclopent-1-en-3-ylamino)-5-(2-(4-iodophenyl)hydrazinyl)-4H-1,2,4-triazole-3-thiol [16] and 4-(cyclopent-1-en-3-ylamino)-5-[2-(4-chlorophenyl)hydrazinyl]-4H-1,2,4-triazole-3-thiol [12]Zarezade et al. [283]2021In silico3D-QSAR pharmacophore modeling ADMET properties, molecular docking molecular dynamic simulation MM-PBSA, hybrid QM-MM method de novo ligand designPubChem and ZINC databasesHuman angiotensin-converting enzyme 2 and main proteaseZINC12562757 and 112,260,215 were the best potential inhibitors of the ACE2 and main protease, respectively. Evo_1 compound enjoys the highest docking energy among the designed de novo ligandsEvo_1 has a stronger potential for specific inhibition of main protease, as compared to the 112, 260, 215 compound. Lead moleculesIn silico studies used the following component of novel coronavirus as targets: main protease (N = 154), spike glycoprotein (N = 62), nonstructural protein (N = 45), RNA-dependent RNA polymerase (N = 21), papain-like protease (N = 19).About 260 drugs were repurposed by the computational methods for COVID-19 therapy such as about 120 drugs candidate against the main protease, 52 drugs against the spike glycoprotein, 14 drugs against RNA-dependent RNA polymerase, and 28 drugs against other nonstructural proteins.Among the studied repurposed drugs, the best results (regarding efficacy) were observed with saquinavir (N = 9 study), ritonavir (N = 8 study), lopinavir (N = 6 study), remdesivir (N = 3 study), and amikacin, danoprevir, favipiravir, and telaprevir.Table1 shows target-based synthesis of data for COVID-19 drug repurposing. As presented, at least two studies show the efficacy of aprepitant, cobicistat, dipyridamole, and dihydroergotamine against the main protease and tegobuvir (N = 2) against spike protein. The following list of drugs had multitarget action: avapritinib, famotidine, bictegravir, ziprasidone, capmatinib, pexidartinib, amprenavir, zafirlukast, cilostazol, paromomycin, lopinavir, and remdesivir.A total of 91 studies used in silico methods to evaluate the effects of natural products including herbal medicine against SARS-CoV-2. Among them, 54 studies used the main protease as main target, in which eucalyptus (N = 3), Tinospora cordifolia (N = 3), and flavonoids (e.g., hesperidin, rutin, and herbacetin) were the most studied and effective. Other studied targets were as follows: spike (N = 22) and multitarget (N = 20). From the plant metabolites, oleanolic acid, hesperidin, epigallocatechin gallate, jensenone, tinosponone, and anistone show promising results in computational methods against COVID-19. Aloe, green tea, eucalyptus, curcumin, and many Chinese, Indian, and African plants were also effective on COVID-19 in silico.The derivatives of pyrazoles, oxadiazoles, phenyltriazolinones, triazoles, benzoylpinostrobin, benzoic acid, benzylidenechromanones, coumarin, and selenium show efficacy in computational methods and could be considered as lead molecules for drug design and synthesis against COVID-19. Melatonin, ebselen, phenyl furoxan, thimerosal, isatin, romidepsin, phenyl mercurin, pleconaril, and tyrosine kinase inhibitors (such as nilotinib and imatinib) are examples of other drugs that are effective in in silico methods.


### 3.2. Results from In Vitro and In Vivo Studies

Screening of in vitro studies leads to finding 34 studies. Different compounds were evaluated and most studies focused on the inhibition of viral replication, which was assessed by the quantification of viral RNA by PCR and IC50 value reported. Most of the studies use the Vero E6 cell line for the assessment of replication of SARS-CoV-2 (Table 4). In particular, main protease, papain-like protease, RNA-dependent RNA polymerase, NSP-14, NSP-15, spike protein, and TMPRSS2 were evaluated in vitro as targets. Also, three studies focused on in vitro inhibition of inflammatory markers such as interleukins and the effects of the cytopathic effect on infected cell.Table 4
Summary of studies with in vitro method.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAgarwal et al. [284]2020In vitroIL-1β assay using THP-1 cells, in vivo pharmacokinetic investigation in C57BL/6 miceAlkenyl sulfonylurea derivativesNLRP3 inhibition and reduction in the release of interleukin-1βIL-1β inhibition IC50 of 35 nM, good oral absorption showing Cmax of 8.49 μg/mL with an AUC of 48.9 μg·h/mL and terminal half-life of 2.86 h, after oral route of administration at 3 mg/kg doseNovel thiazolo-alkenyl sulfonylurea derivative7 as potent, selective, and orally bioavailable NLRP3 inflammasome inhibitorAkaberi et al. [285]2020In vitroVero E6 cells, RT-qPCR for the quantification of viral RNA, expression and purification of SARS-CoV-2 3CL protease in vitro enzymatic assayNO-Donor S-nitroso-N-acetylpenicillamine (SNAP): nitric oxideMain proteaseAlthough the viral replication was not completely abolished (at 200μM and 400 μM), SNAP delayed or completely prevented the development of viral cytopathic effect in treated cells (IC50 = 440.95 μM ± 36.15 SE)A dose-dependent inhibition of the 3CL protease (400μM)Bernstein and Zhang [286]2020In vitroVero E6 cells: virus was then added at a multiplicity of infection (MOI) of 0.01, with 1 h allowed for infection. Cytotoxicity: Cell Counting Kit-8 (CCK-8) colorimetric assayGallium maltolate (GaM)Inhibition of viral replicationEC50 (concentration producing 50% inhibition of viral replication) of about 14 μM (CI 95%: 8.9–22.8 μM)No cytotoxicity was observed at concentrations up to at least 200μMBocci et al. [287]2020In vitroLigand-based virtual screening/Vero E6 cells. Uninfected cells and chloroquine as control inhibition of the SARS-CoV-2-mediated CPE following infection in Vero E6 cells tissue culture infectious dose (TCID50) assay4000 approved drugs/glafenine, amodiaquine vorinostat, zuclopenthixol, isoxsuprine, nebivolol, ambroxol, panobinostat, and pracinostatInhibition of viral replicationIn silico: glafenine, amodiaquine (amodiaquine), vorinostat, zuclopenthixol, isoxsuprine, nebivolol, ambroxol, panobinostat, and pracinostat. In vitro: zuclopenthixol and amodiaquine show anti-SARS-CoV-2 activity comparable with chloroquine. EC50 values were estimated as 0.13 μM for AQ, 1.35 μM for ZPX, and 2.72 μM for nebivololZuclopenthixol and nebivolol exhibitin vitro antiviral activities and potencies that are comparable or better than chloroquine and hydroxychloroquineCao et al. [288]2020In vitroAfrican green monkey kidney Vero E6 cells were infected with virus at an MOI of 0.01 for 1 h. Immunofluorescence assay physiologically based pharmacokinetic modeling and simulations9 artemisinin-related compoundsInhibition of viral replicationArteannuin B showed the highest anti-SARS-CoV-2 potential with an EC (50) of 10.28 ± 1.12μM. Artesunate and dihydroartemisinin showed similar EC (50) values of 12.98 ± 5.30 μM and 13.31 ± 1.24 μM, respectivelyLumefantrine could inhibit SARS-CoV-2in vitro with an EC50 of 23.17 ± 3.22 μM. Impair viral infection by modulating host cell metabolic pathwaysChen et al. [289]2021In vitroFluorescence-based high-throughput screen, docking/pretreated with various concentrations of test compound for 1 h, followed by infection with SARS-CoV-2 (MOI of 0.0001) in the presence of test compounds, expression and purification of SARS-CoV-2 PLpro, 3CL protease fromEscherichia coli enzymatic assay of SARS-CoV-2 PLpro cytotoxicity assay1920 natural productsMain protease, papain-like proteaseGinkgolic acid and anacardic acid were also identified as inhibitors of 3CLpro, with IC50 values of 1.79 ± 0.58 and 2.07 ± 0.35 μM, respectively, both ginkgolic acid and anacardic acid dose-dependently inhibited PLpro activity, with IC50 values of 16.30 ± 0.64 and 17.08 ± 1.30 μM, respectivelyGinkgolic acid act as an irreversible inhibitor against both PLpro and 3CLpro, suggesting it is a covalent inhibitorGendrot et al. [290]2020In vitroVero E6 cells, replication was estimated by RT-PCR/chloroquine as control determination of the inhibition stageDoxycyclineInhibition of viral replicationMedian effective concentration (EC50) and 90% effective concentration (EC90) for doxycycline were 4.5 ± 2.9 μM and 23.5 ± 16.5 μM, respectively, EC50 = 4.5 μMInteracted at both entry and postentry stages of the SARS-CoV-2 infection doxycycline has anti-inflammatory effectsGupta et al. [291]2021In vitroHigh-throughput virtual screening, molecular dynamic simulation, prime MM-GBSA/Vero E6 cells infected with the SARS-CoV-2 fluorescence-based biochemical assay for inhibitors of 3CLproFDA-approved drugsMain protease, PLpro, RNA-directed RNA polymerase, helicase, NSP-15, NSP14, and 2′-O-methyltransferaseTroxerutin (main protease and PLpro) and bisindolylmaleimide derivatives (main protease and exon) had possible dual targets. Ivermectin has nonselective toxicity to the ATCC E6 Vero cells at ≤50μM and 16.67 μM based on the number of nuclei countedBIM IX specifically blocked 3CLproin vitro enzymatic assay inhibition was observed with an IC50 value of 113.7 ± 5.2 μM a known inhibitor of protein kinase C isoforms, bisindolylmaleimide IX (BIM IX), was found to be a potent inhibitor of SARS-CoV-2Hahn et al. [292]2020In vitroVero B4 and 76 cells و Vero E6 cells/remdesivir RT-qPCR for the detection of extracellular SARS-CoV-2 in cell immunostaining for the detection of Intracellular SARS-CoV-2 Western blot analysis viral plaque and yield reduction assay neutral red assay (NRA)(IMU-838) inhibitor of human dihydroorotate dehydrogenase (DHODH)Inhibition of viral replication/immunomodulatorA virus-specific EC90 of 6.2 ± 1.9 μM was measured, with no cytotoxicity observed with drug concentrations up to 100 μM at 3 days p.i.Active moiety of IMU-838, vidofludimus, possesses broad-spectrum antiviral activity. IMU-838 reduces T lymphocyte proliferation, cytokine production, and organ infiltration by leukocytes in various in vivo and in vitro models for autoimmunityHattori et al. [293]2021In vitroVeroE6 cell-based assays with RNA-qPCR, cytopathic assaysTwo small-molecule-compounds, named GRL-1720 and 5 h, containing an indoline and indole moietyMain proteaseEC(50) values of 15 ± 4 and 4.2 ± 0.7μM for GRL-1720 and 5 hCombination of 5 h and remdesivir exhibits synergism against SARS-CoV-2.5 h might serve as a lead M (pro) inhibitorJang et al. [294]2021In vitroVero CCL-81 cells/Calu-3 human lung epithelial cellsGemcitabine and its analog 2′-fluoro-2′-deoxycytidine (2FdC)Inhibition of viral replication50% effective concentration (EC(50)) of 1.2μM compare to remdesivir 2FdC was marginally active (EC(50) = 175.2 μM)Gemcitabine has a synergistic effect when combined with remdesivirJeon et al. [295]2020In vitroDrug repositioning/chloroquine, lopinavir, and remdesivir were used as reference drugs/VeroE6 cell/immunofluorescence analysis48 FDA-approved drugsInhibition of viral replicationNiclosamide (IC50, 0.28 μM), ciclesonide (IC50, 4.33 μM)24 potential antiviral drug candidates against SARS-CoV-2 infectionLiu et al. [296]2020In vitroVirtual screening/spiked pseudotyped lentivirus for imitating SARS-CoV-2 cell entry/ACE2-GFP expressing HEK293T 2 cells. HeLa Cells, 293T (human, kidney) cells, and Vero-E6 (African green monkey, kidney) cellsFDA-approved drugsAngiotensin I-converting enzyme 2 (ACE2) and a spike proteinNine potential candidates were selected and submitted to experimental studies. Three (romidepsin, saquinavir, and nelfinavir) of nine drugs possess the ability to suppress SARS-2-S pseudotyped particles to enter the ACE2 expressing cells in a concentration-dependent mannerFive clinical HDAC inhibitors including romidepsin, panobinostat, givinostat hydrochloride monohydrate, CAY10603, and sirtinol could inhibit noticeably the spike/ACE2-mediated cell entry of SARS-CoV-2Ma et al. [297]2020In vitroVero E6 cells using cytopathic effect and plaque reduction assay. HCoV-229E infection in Huh-7 cells. Western blot assayPhillyrin (KD-1) ingredient of Forsythia suspensaInhibition of viral replicationInhibit SARS-CoV-2. Markedly reduce the production of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β, MCP-1, and IP-10) at the mRNA levels. p-NF-κB p65, NF-κB p65, and p-IκBαKD-1 could significantly inhibit virus proliferation in vitro, the upregulated expression of pro-inflammatory cytokines induced by SARS-CoV-2Masih et al. [298]2020In vitroLPS-stimulated RAW267.4 cells by enzyme immunoassay Western blot assayPyrazole derivativesAnti-inflammatory levels of interleukin-1β, tumor necrosis factor-α, and interleukin-6Compound 6c as the most potent analog among the tested series. Inhibition of inhibitor kappa B-α and NF-κBOmotuyi et al. [299]2020In vitroKinetic assayAframomum melegueta. 100 secondary metabolitesMain protease, 2′-O-ribose methyltransferase (NSP16), and surface glycoprotein/ACE2 receptor interfaceDiarylheptanoid (letestuianin A), phenylpropanoid (4-cinnamoyl-3-hydroxy-spiro[furan-5,2′-(1′H)-indene]-1′,2,3′(2′H,5H)-trione), flavonoids (quercetin, apigenin, and tectochrysin) have been identified as high binding compounds to SARS-CoV-2 targets in a polypharmacology mannerDi-ethyl-ether (IC50 = 0.03 mg/L), acetone (IC50 = 1.564 mg/L), ethyl-acetate (IC50 = 0.382 mg/L), and methanol (IC50 = 0.438 mg/L) fractions demonstrated the best inhibition in kinetic assayOutlaw et al. [300]2020In vitroVero E6 cellβ-Gal complementation-based fusion assay. Viral titration and plaque reduction neutralization assay quantitative RT-PCR.A lipopeptide that is derived from the C-terminal heptad repeat (HRC) domain of SARS-CoV-2 SInhibition cell-cell fusionInhibits cell-cell fusion mediated by SARS-CoV-2 S and blocks infection by live SARS-CoV-2 in Vero E6 cell monolayers (IC50) of ∼10 nM and 90% inhibitory concentration (IC90) of ∼100 nMPitsillou et al. [301]2021In vitroMolecular docking, molecular dynamic simulation/papain-like protease (SARS-CoV-2) assay kit GRL-0617 as a control enzymatic inhibition assay300 small compounds derived predominantly from our OliveNet™ library (222 phenolics) and supplemented with synthetic and dietary compoundsPapain-like proteaseSelection of 30 compounds. Hypericin possessed inhibition activity, and both rutin and cyanidin-3-O-glucoside resulted in a concentration-dependent inhibition of the PLproThe IC50 values were not clarified in experimentPizzorno et al. [302]2020In vitroVero E6 cells/remdesivir, lopinavir, chloroquine as comparatorFavipiravir, ribavirin, umifenovir (arbidol), berberine, cyclosporine A, diltiazemInhibition of viral replicationUmifenovir, berberine, and cyclosporine A with estimated 50% inhibitory concentrations of 0.99, 5.2, 1.38, 3.5, 10.6, and 3μM, respectivelyA strong antagonism between remdesivir and berberine, in contrast with remdesivir/diltiazem, for which we describe high levels of synergyPlaze et al. [303]2021In vitroMonkey VeroE6 cells and human alveolar basal epithelial A549-ACE2 cell RT-qPCR for the presence of SARS-CoV-2 RNAChlorpromazineInhibition of viral replication(IC50) of 8.2 μM in monkey VeroE6 cells (CC50) of 13.5 μM, and selectivity index (SI) of 1.65 IC50 of 11.3 μM, CC50 of 23.1 μM, and SI of 2.04 in human A549-ACE2 cellsReznikov et al. [304]2021In vitroRepurposing study molecular docking/human lung A549 cells that were transfected with hACE2 (ACE2-A549) cells, Vero E6 cellsElectronic health records of over 219,000 patients with antihistaminesInhibition of viral replication/spike proteinIC50 15.3 μg/ml for hydroxyzine, 17.4 μg/ml for diphenhydramine, and 2.24 μg/ml for azelastineHydroxyzine and possibly azelastine bind angiotensin-converting enzyme-2 (ACE2) and the sigma-1 receptor as off-target disease preventionSakurai et al. [305]2021In vitroVeroE6 cells/human colon-derived Caco-2 cells/infection assay with immunofluorescence5-Amino levulinic acid (5-ALA)Inhibition of viral replicationIC50 of 570 μM/IC50 of 63 μM in human Caco-2 cells. No significant cytotoxicity5-ALA is synthesized in most animals and plants and we are continuously consuming it in our food. Show anti-inflammation effects in humansStone et al. [306]2021In vitroVero E6 monkey kidney and Calu-3 human lung adenocarcinoma cells/RT-qPCR of RNAStenoparib poly(ADP ribose) polymerase (PARP) inhibitorInhibition of viral replicationDose-dependent activity against SARS-CoV-2 at concentrations up to 30μM with negligible cytotoxicity. Synergistic effect with remdesivirStenoparib impedes entry and postentry processesSu et al. [307]2020In vitroVero E6 cells, quantitative real-time RT-PCRShuanghuanglian preparation, a Chinese traditional patent medicine (chlorogenic acid, phillyrin, baicalin, and baicalein)Main proteaseDose-dependent inhibition of SARS-CoV-2 3CLpro, and the resulting IC50 values were 0.090, 0.064, and 0.076 μL/mL for three Shuanghuanglian oral liquids produced by three different pharmaceutical companiesBaicalin and baicalein, which have good drug-like propertiesTouret et al. [308]2020In vitroVeroE6 cells/human colon-derived Caco-2 cell real-time RT-PCR/remdesivir as controlFDA-approved chemical libraryInhibition of viral replication23 drugs were selected to cover the 12 different groups. 11 compounds such as macrolide antibiotics, proton pump inhibitors, antiarrhythmic agents, or CNS drugs emerged showing antiviral potency with 2 < EC50 ≤ 20μMTwo of the highest antiviral activity were obtained for azithromycin (EC50 = 2.12μM) and hydroxychloroquine (EC50 = 4.17 μM). Vonoprazan and sulfadoxine show a moderate activity with EC50 above 30 μMTree et al. [309]2021In vitroPlaque inhibition assay with Vero E6 cells/differential scanning fluorimetry/ELISA assaysUnfractionated heparin (UFH), low MW heparinsInhibition of viral replication/spike protein RBDAll the UFH preparations had potent antiviral effects, with IC50 values ranging between 25 and 41 μg·ml−1, whereas LMWHs were less inhibitory by ∼150-fold (IC50 range 3.4–7.8 mg·ml−1)Heparin directly inhibits the binding of RBD to the human ACE2 protein receptorVatansever et al. [310]2021In vitroDocking, screening assay/Vero E6, and A549 cell culture30 FDA/EMA-approved drugsMain proteaseSix FDA/EMA-approved drugs can potently inhibit Mpro with an IC50 value lower than 100 μM. Bepridil exhibited strong inhibition of SARS-CoV-2 from entry and replication inside Vero E6 and A549 cells (IC50 = 72 μM)Bepridil indicated that it had low micromolar EC50 valuesWang et al. [311]2020In vitroFRET-based enzyme activity assay of Mpro measurement of human TMPRSS2 activity by a FRET-based enzymatic assay surface plasmon resonance (SPR) analysis molecular docking African green monkey kidney cell Vero E6Catechin, kaempferol, quercetin, proanthocyanidins, resveratrol, and tannic acidTMPRSS2 (transmembrane protease serine 2) main proteaseTannic acid inhibited the activities of the two proteases with an IC50 of 13.4 mM for Mpro and 2.31 mM for TMPRSS2A potent dual inhibitorWatashi [312]2021In vitroVirtual drug screening, docking/VeroE6 cells detecting the viral RNA by real-time RT-PCR or viral proteins by immunofluorescenceFDA-approved drugsMain protease, PLpro, RdRp, helicase, spike, ACE2, and TMPRSS2Abiraterone, amodiaquine, anidulafungin, arbidol, astemizole, atazanavir, auranofin, azithromycin bazedoxifene, bexarotene, camostat IC50 values <5 μMCepharanthine, cetilistat, chloroquine, ciclesonide, cyclosporine A, digoxin, ivermectin, lopinavir, mefloquine, nelfinavir, niclosamideYu et al. [313]2021In vitroComputer-aided drug design and biological verification surface plasmon resonance (SPR) assays and NanoBiT assaysTraditional Chinese medicineSpike proteinsGlycyrrhizic acid, the most efficient and nontoxic broad-spectrum anticoronavirus molecule IC50, was 22 μMZandi et al. [314]2021In vitroAfrican green monkey kidney cells (Vero CCL-81 cells) and reverse transcription-quantitative PCR (qRT-PCR). Remdesivir andβ-D-N4-hydroxycytidine as positive drug controlsNucleoside analogs/anti-HCV agentsInhibition of viral replicationSofosbuvir and favipiravir demonstrated no antiviral effect against COVID-2Zhang et al. [315]2020In vitroHybrid virtual screening, docking, force field-based simulation/qRT-PCR assay, indirect immunofluorescence assay (IFA) and CCK-8 assay, surface plasmon resonance (SPR) assay/Vero cells1906 approved drugs from TargetMol-approved drug libraryRNA-dependent RNA polymerase (RdRp)Pralatrexate and azithromycin (EC50) values of 0.008 and 9.453 μMA new therapeutic agent pralatrexate against COVID-19 by targeting RdRpZhang et al. [316]2020In vitroAfrican green monkey kidney Vero E6 cell line, quantitative real-time PCR (RT-PCR) analysis, immunofluorescence microscopy, Western blot analysisLopinavir/ritonavir, rupintrivir, and AG7404Inhibition of viral replication(IC50) and half cytotoxic concentration (CC50) values of lopinavir were ∼12.01 μmol/L and 80.82 μmol/L; for ritonavir, the IC50 and CC50 values were 19.88 μmol/L and 94.71 μmol/L (SI = 4.76)Rupintrivir inhibited SARS-CoV-2 infection only at high drug concentrations (Vero E6: IC50 = 34.08 μmol/L, AG7404: Vero E6: IC50∼195.8 μmol/L). Lopinavir/ritonavir should be stopped for clinical use due to the huge gap between in vitro IC50 and free plasma concentrationZhu et al. [317]2021In vitroHuman fibroblast lung cells (MRC-5), cytopathic effect (CPE) assay, primary human airway air-liquid interface (ALI) cultures, TEER assay, CCK-8 assaysStimulator of interferon genes (STING): dimeric amidobenzimidazole (diABZI)Inhibition of viral replicationPotent anticoronavirus activity against both the common cold human coronavirus 229E (HCoV-229E) and SARS-CoV-2 in cell culture systems. EC50 of 120 μMNitric oxide, ginkgolic acid, anacardic acid, troxerutin, bisindolylmaleimide derivatives, small molecules (GRL-172, and 5 h), baicalin and baicalein (phytochemicals), and bepridil were effective drugs in vitro against the main protease of COVID-19.The following drugs could inhibit the spike/ACE2-mediated cell entry of COVID-19 in vitro: romidepsin, panobinostat, givinostat, sirtinol, saquinavir, lipopeptides, hydroxyzine, azelastine, heparin, and glycyrrhizic acid.Alkenyl sulfonylurea derivatives [7], an IMU-8381 inhibitor of human dihydroorotate dehydrogenase, pyrazole derivatives, and phillyrin, regulated the expression of inflammatory cytokines (e.g., IL, TNF-α, and NF-κB) induced by SARS-CoV-2 markedly and could be considered as adjuvant treatment of COVID-19 severe disease.Doxycycline, chlorpromazine, azithromycin, heparin, bepridil, tannic acid, and glycyrrhizic acid are well-known drugs that show both in silico and in vitro inhibitory effects against SARS-CoV-2 and should be considered for this purpose.Also, in vitro studies show that lopinavir/ritonavir, sofosbuvir, and favipiravir have no antiviral effects against SARS-CoV-2 (huge gap between in vitro IC50 and free plasma concentration).We included 16 in vivo studies in our final analysis. Most of them use a combination of in vitro and in vivo methods for the evaluation of novel drugs. Table5 shows the detail of these studies.Table 5
Summary of studies with in vivo method.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsLei et al. [318]2020In vivoBALB/c mice to determine the pharmacokinetic profiles of the fusion proteinsA recombinant protein by connecting the extracellular domain of human ACE2 to the Fc region of the human immunoglobulin IgG1Spike proteinThe IC50 values of ACE2-Ig for SARS-CoV and SARS-CoV-2 neutralization were 0.8 and 0.1 μg·ml−1, respectively. Desirable pharmacological properties in micePotential applications in the diagnosis, prophylaxis, and treatment of SARS-CoV-2Rathnayake et al. [319]2020In vivoOld male hDPP4-KI mice infected with MERSMA-CoVDipeptidyl and tripeptidyl series of compoundsMain protease7a, 6c, 7e, 7h, and 6j: EC50 values ranging from 0.15 to 0.9 μM in Vero E6 cells, IC50: 0.17 to 0.82 μM. 40% of mice treated with compound 6h survived, and all mice treated with compound 6j were alive at the end of the study6j resulted in the survival of MERSMA-CoV-infected hDPP4-KI miceSchäfer et al. [320]2020In vivoAn immune-competent mouse model of COVID-19 by remodeling the SARS-CoV-2 S RBD at the mACE2-binding interface. Human Ace2-expressing HT1080 cells and Syrian hamster model of SARS-CoV-2Human mAbs (hu-mAbs)Reduction in viral load and prevent infectionNeutralizing hu-mAbs targeting SARS-CoV-2 promotes reduction in viral load and prevent infection in macaques and hamsters effective dose: 5.3 and 16 mg/kg inhibitory concentration (IC)50/IC90 (half-maximal/90% IC) of 4.4/18 to 26/140 ng/mlSchumann et al. [321]2020In vivoFemale C57Bl/6 mice, LPS stimulationMP1032:5-amino-2,3-dihydro-1,4-phthalazinedione sodium saltImmune-modulatingDirect inhibitory activity on human PARP-1 was detected in the low micromolar range (IC50 = 1.55 μM). MP1032 works as a ROS scavenger, reduced the secretion of TNF-α and IL-6. MP1032 pre-treatment significantly reduced plasma cytokine levels compared with vehicle-treated mice by 50% (TNF-α) and 25% (IL-6). Significant reduction in SARS-CoV-2 replicationMP1032 could further attenuate prolonged virus replication by preventing oxidative stress or by limiting ADP ribosylation of the viral nucleocapsid protein via PARP-1 inhibitionShang et al. [322]2021In vivoAfrican green female hACE2 transgenic mice, SARS-CoV-2 injection RNA extraction and qPCR, RNA in situ hybridization assay, histopathological examinationEndosomal acidification inhibitors, including chloroquine, bafilomycin A1, and NH4CLAntiviral actions against SARS-CoV-2(40μM), bafilomycin A1 (100 nM), and NH4CL (12.5 mM) suppressed the replication of SARS-CoV-2 in all cell types chloroquine (60 mg·kg−1) and bafilomycin A1 (0.1 mg·kg−1) markedly reduced virus yields in lung tissues. Alleviated viral pneumonia in hACE2 transgenic miceEndosomal acidification inhibitors exhibited antiviral actions against SARS-CoV-2Sheahan et al. [323]2020In vivoMale and female 20- to 29-week old SPF C57BL/6J (Stock 000664 Jackson Labs) mice, whole-body plethysmography 50, 150, or 500 mg/kg EIDD-2801 2 hr prior to intranasal infection with 5E + 04 PFU of mouse-adapted SARS-CoVRibonucleoside analogβ-d-N4-hydroxycytidineAntiviral efficacy against the three most recently emerged human CoV: SARS-CoV, MERS-CoV, and SARS-CoV-2Bioavailable NHC prodrug (β-d-N4-hydroxycytidine-5′-isopropyl ester), improved pulmonary function and reduced virus titer and body weight lossContinued development of EIDD-2801 as a potent broad-spectrum antiviralTai et al. [324]2021In vivoSprague Dawley rats. (i) HCQ‐IV: 12 rats received a single dose of 0.590 mg HCQ sulfate per animal via IV injection; (ii) HCQ‐IT: 20 rats received a single dose of 0.590 mg HCQ sulfate per animal via intratracheal (IT) administration; and (iii) liposomal HCQ‐IT: 20 rats received a single dose of 0.284 mg liposomal HCQ sulfate per animal via IT administration 0.25, 1, 4, 24, and 72 hours postdose and for tissue/organ samples were 0.25, 4, 24, and 72 hours postdoseInhalable liposomal hydroxychloroquine (HCQ)Pharmacokinetic studyCompared with unformulated HCQ administered intravenously, liposomal HCQ showed higher (∼30-fold) lung exposure, longer (∼2.5-fold) half-life in lungs, but lower blood exposure with ∼20% of peak plasma concentration (Cmax) and 74% of area under the curve from 0 to 72 hours (AUC 0–72) and lower heart exposure with 23% of Cmax and 58% of AUC 0–24 (normalized for dose)Inhalable liposomal HCQ may provide clinical benefit and serve as a potential treatment of COVID-19Tortorici et al. [325]2020In vivoFrom two individuals recovering from severe COVID-19 disease. Surface plasmon resonance (SPR) and flow cytometry. Cell-cell fusion assay using VeroE6 cells/Syrian hamster modelHuman neutralizing antibodies (S2E12 and S2M11)Against SARS-CoV-2IC50 values were 1.2 to 6.6 ng/ml to protect hamsters against SARS-CoV-2 challenge competitively block angiotensin-converting enzyme 2 (ACE2) attachment and that S2M11 also locks the spike in a closed conformation by recognition of a quaternary epitope spanning two adjacent receptor-binding domainsAntibody cocktails for prophylaxis or therapyWahl et al. [326]2021In vivoCoronavirus replication in human lung-only mice (LoM) and histopathologic analysis LoM were administered EIDD-2801 starting 24 h or 48 h post-SARS-CoV-2 exposure and every 12 h thereafterRibonucleoside analogβ-D-N4-hydroxycytidine (NHC): oral prodrug EIDD-2801 (also known as molnupiravir or MK-4482)Inhibited SARS-CoV-2 replicationEIDD-2801 pre-exposure prophylaxis significantly reduced virus titers in the human lung tissues of LoM by over 100,000-fold in two independent experimentsProphylactic administration EIDD-2801 is highly effectiveWang et al. [327]2021In vivoMale hACE2 mice inoculated intranasally with SARS-CoV-2 stock virus rhesus macaque model of SARS-CoV-2 infectionScreening the FDA-approved peptide drug library with LibDockAgainst SARS-CoV-2Ten polypeptide drugs were selected. Dalbavancin showed the strongest inhibitory ability. (EC50) was ∼12 nM. Significant inhibition of SARS-CoV-2 pseudovirion entry into HEK293/hACE2 cells, with an IC50 of ∼53 nM in both mouse and rhesus macaque models, viral replication, and histopathological injuries caused by SARS-CoV-2 infection are significantly inhibited by dalbavancin administrationDalbavancin directly binds to human angiotensin-converting enzyme 2 (ACE2) with high affinity, thereby blocking its interaction with the SARS-CoV-2 spike proteinWeston et al. [328]2020In vivoMice were inoculated intranasally with SARS-CoV20 FDA-approved drugsAgainst SARS-CoV-27 of these inhibit SARS-CoV-2 at non-cytotoxic concentrations. Many of the compounds have IC50 under 10 μM, chloroquine and chlorpromazine did not inhibit viral replication in mouse lungs based on viral titers recovered at 4 dpi neither drug inhibited viral replication in the lungs, but both protected against clinical diseaseWhite et al. [329]2021In vivoA human ACE2-expressing adenovirus to transduce the naturally resistant wild-type BALB/c mice and sensitize them to SARS-CoV-2 infection, K18-hACE2 mouse modeleEF1A inhibitor plitidepsin (aplidin): targeted the eukaryotic translation machineryInhibited SARS-CoV-2 replicationIC90 of 1.76 nM: Vero E6 IC90 of 0.88 Nm: human cell line significantly reduced genomic RNA content reduction of nearly 2 log units in SARS-CoV-2 viral titers in the lungs of the 0.3 mg/kg plitidepsin group, reduction of 2 log units in viral lung titers at day 3, similar to two daily 50 mg/kg doses of remdesivirPlitidepsin as a host-targeted anti-SARS-CoV-2 agent with in vivo efficacyWu et al. [330]2020In vivo10 male C57BL/6 mice related to 200 mg/kg/day GB-2 everyday by oral administration. immunohistochemistry (IHC) assessmentGB-2, the formula from Tian Shang Sheng Mu of Chiayi Puzi Peitian Temple: in Taiwan: the index compound ofCamellia sinensis var. assamica extractThe protein and mRNA expression of ACE2 and TMPRSS2No mouse death and no significant alteration in mouse activity or body weight (IHC) data revealed that the expression levels of both ACE2 and TMPRSS2 were markedly diminished in the GB-2 group compared with the control group 50μg/mL of theaflavin could inhibit protein expression of ACE2 and TMPRSS2GB-2 and theaflavin could act as potential compounds for ACE2 and TMPRSS2 inhibitorsYuen et al. [331]2021Ex vivoHuman lung tissues were treated with the indicated drugs overnight, followed by SARS‐CoV‐2 infection at 2 × 106 PFU/wellSmall molecules targeting the ULK1/Atg1 complex involved in the induction stage of autophagy (ULK1 inhibitor SBI0206965), the ATG14/Beclin1/VPS34 complex involved in the nucleation step of autophagy (class III PI3‐kinase inhibitor VPS34‐IN1), and a widely used autophagy inhibitor that persistently inhibits class I and temporary inhibits class III PI3‐kinase (3‐MA) and a clinically approved autophagy inhibitor that suppresses autophagy by inhibiting lysosomal acidification and prevents the formation of autophagolysosome (HCQ)Target the key cellular factors involved in key steps of the autophagy pathwayNot all the tested autophagy inhibitors suppressed SARS-CoV-2 infection inhibition of class III PI3‐kinase Vps34 downstream of ULK1, in contrast to inhibition of ULK1, which caused a significant decrease in SARS‐CoV‐2 replication (EC50) of HCQ was 19μM, that of Vps34‐IN1 was 0.82 μM Vps34‐IN1 potently inhibited SARS‐CoV‐2 viral replication in normal ex vivo human lung tissue culture in a dose‐dependent mannerClass III PI3-kinase may be a possible target for COVID-19 therapeutic developmentZhang et al. [332]2021In vitro/in vivoPreparation of MAbs from female BALB/c mice and wild-type Balb/c mice were intranasally inoculated with hACE2-encoded adenovirus 5 (Ad5-hACE2) to allow expression of the hACE2 receptor in the lung, followed by intranasal infection with live SARS-CoV-2 3 days laterMouse anti-SARS-CoV-2-neutralizing MAbs: 2H2 and 3C1Spike (S) protein(IC50) of 12 ng/mL effectively treat SARS-CoV-2-infected mice even when administered as late as 24 h post-infectionHuman monoclonal antibodies were the most evaluated drugs (N = 4), promote the reduction in viral load (in vitro), and prevent infection in animal models of SARS-CoV-2.A prodrug of hydroxycytidine (molnupiravir) improved pulmonary function and reduced viral titer in vivo and was introduced as a potential broad-spectrum antiviral agent against SARS-CoV-2.Chloroquine and chlorpromazine did not inhibit viral replication in mouse lungs, but protected them against clinical disease.Dalbavancin shows significant inhibitory ability in both in vitro and in vivo models of COVID-19. The drug binds directly to ACE2 and blocks its interaction with the SARS-CoV-2 spike protein.


## 3.1. Results from In Silico Drug Discovery

From 267 studies used in silico approaches, 98 article studies repurposed approved drugs with a new mechanism of action and 91 studies evaluated natural products (e.g., herbal medicine) on COVID-19. The characteristics of these studies are summarized in Tables1 and 2. Also, Table 3 shows the characteristics of the remaining studies (N = 87).Table 1
Summary of studies with in silico method that used repurposed drugs for novel drug discovery against COVID-19.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAbosheasha and El-Gowily [24]2020In silicoDrug repurposing molecular docking-based virtual screening15 antiplatelet FDA-approved drugsMain protease (Mpro) and spike glycoprotein (S)Cilostazol has favorable binding interaction with Mpro (PDB ID: 6LU7) cilostazol, iloprost, epoprostenol, prasugrel, and icosapent ethyl that have a higher binding affinity on spike glycoprotein (S)Cilostazol is a promising FDA drug against COVID-19 by inhibiting both Mpro and S proteinAbdul Kadhim et al. [25]2020In silicoDrug repurposing, dockingExperimental and approved drugsPapain-like protease and RNA polymeraseDrugs that shared >70% similarity to the binding sites of those targets were reversin, pentagastrin, remdesivir, norfloxacin, and nitazoxanide against COVID-19 papain-like protease whereas benzylglutathione, lopinavir, and hydroxymethylglutathione against RNA polymeraseAntiresistance reversin showed the highest inhibitory efficacy against COVID-19 papain-like protease, and benzylglutathione is an experimental compound; however, it had the highest RNA polymerase inhibiting efficacyAbu-Saleh et al. [26]2020In silicoLigand-based/structure-based virtual screening, MD simulations, and binding energy calculationsApproved drugs and bioactive compounds listed in the DrugBank and ChEMBL databasesMain proteaseBest MM/GBSA binding energy; ChEMBL275592, montelukast, ChEMBL28834. Bromocriptine and saquinavir demonstrate stability in the active site of MproAchilonu et al. [27]2020In silicoHigh-throughput virtual screening and ligand dockingFDA-approved drugsMain proteaseIsavuconazonium, a P2-P3α-ketoamide derivative, and pentagastrina are the top three molecules (Lig13b as the benchmark) based on docking energyAftab et al. [28]2020In silicoRepositioning/target-based virtual screening and molecular dockingTen antiviral drugs were screened: ribavirin, remdesivir, sofosbuvir, penciclovir, nitazoxanide, nafamostat, chloroquine, galidesivir, favipiravir, and interferonRNA-dependent RNA polymerase (RdRp)Galidesivir and its drug-like compounds CID123624208 and CID11687749 have shown an effective attachment to the priming site of viral RdRpCID123624208 and CID11687749 may be considered for in vitro and in vivo clinical trialsAhmadi et al. [29]2021In silicoDrug repurposing study using molecular dockingEnfuvirtide, an HIV-1 fusion inhibitor peptideSARS-CoV-2 fusion inhibitorEnfuvirtide binding to the S2 protein of SARS-CoV-2 was remarkably stable and can act as a strong SARS-CoV-2 fusion inhibitorAhmed et al. [30]2021In silicoDrug repurposing (high-throughput virtual screening) (HTVS) followed by re-docking with standard precision (SP) and extra-precision (XP) molecular dockingFDA-approved antiviral and anti-infection drugsMain proteaseOf 1397 potential drugs, 157 showed considerable affinity towards Mpro. High-affinity lead drugs (iodixanol, amikacin, troxerutin, and rutin) were identified. Amikacin was found to be the most potent inhibitor of main proteaseAminoglycosides may serve as a scaffold to design potent drug molecules against COVID-19Anand et al. [31]2021In silicoMolecular docking130 US FDA-approved drugs including hypertension, cardiovascular diseases, respiratory tract infections (RTI), antibiotics, and antiviral drugsStructural and nonstructural proteins of SARS-CoV-2 (nsp3, nsp5, nsp10, nsp16)15 potent drugs exhibiting significant inhibitory potential against SARS-CoV-2 like baloxavir marboxil, danoprevir and sofosbuvir, fosinopril, moexipril, quinapril, telmisartan, azilsartan, verapamil, and doxazosinAzithromycin, doxycycline, clarithromycin, rifamycin, and augmentin: nsp10; virginiamycin, tunicamycin, quinupristin, fidaxomicin, digoxin, and azithromycin: main protease; caspofungin, amphotericin B, ketoconazole, and micafungin: E and N proteins; virginiamycin and amphotericin B: S proteinAncy et al. [32]2020In silicoMolecular docking, molecular dynamics, and binding free energy simulation studyHIV-1 protease, namely, TMB607 and TMC-310911Main proteaseTMB607 molecule binds strongly with the SARS-CoV-2 main protease enzymeAnsari et al. [33]2020In silicoRepurposing drug molecular dockingTAT-peptide 47–57 (GRKKRRQRRRP)-conjugated repurposed drugs (i.e., lopinavir, ritonavir, favipiravir, and hydroxychloroquine)Main proteaseTP-conjugated ritonavir, lopinavir, favipiravir, and hydroxychloroquine have superior and significantly enhanced interactions with main proteaseArun et al. [34]2020In silicoRepurposing drug molecular dockingDrugs available in the super DRUG2 databaseMain proteaseBinifibrate and bamifylline bind strongly to the enzyme active siteArya et al. [35]2020In silicoMolecular dockingFDA-approved drugsPapain-like protease15 FDA-approved drugs, including chloroquine and formoterol, bind the target enzyme with significant affinity and good geometry, suggesting their potential to be utilized against the virusBaby et al. [36]2020In silicoSchrodinger’s computer-aided drug discovery tools for in silico drug repurposingFDA-approved library of drugsRNA-dependent RNA polymerase (RdRp)Pitavastatin, ridogrel, rosoxacinBaby et al. [36]2021In silicoSchrodinger’s computer-aided drug discovery tools for in silico drug repurposingFDA-approved library of drugsMain proteaseTipiracil and aprepitant interacted with the main proteaseBaker et al. [37]2021In silicoRepurposing drug molecular docking50 compounds with activity against main proteaseMain proteaseDrugs including boceprevir, ciluprevir. narlaprevir, and telaprevir may be more potent against main protease than boceprevir and suitable for rapid repurposingBharath et al. [38]2020In silicoDrug repurposing computer-aided drug design (CADD)4015 known and approved small moleculesSpike glycoproteinGlycyrrhizic acid (GA) of plant origin may be repurposed for SARS-CoV-2 interventionBhowmik et al. [39]2021In silicoRepurposing drugs, docking, and molecular dynamic simulationOrientin (phytochemical)Inhibitor of SARS-CoV-2 spike and host cell receptor GRP78 bindingBinding of orientin in the overlapping residues of GRP78 binding region of SARS-CoV-2 spike modelAs a promising precautionary or therapeutic measure for COVID-19Bolelli et al. [40]2021In silicoDrug repurposing, virtual screening methodFDA-approved drugsMain proteaseThree compounds (dobutamine and its two derivatives)Cavasotto et al. [41]2021In silicoDrug repurposing, docking-based screening using a quantum mechanical scoringFDA-approved drugsSpike protein, main protease papain-like proteaseSovaprevir, elbasvir, danoprevir, samatasvir, Candesartan, saquinavir ritonavir, indinavir, lopinavir, brilacidin, flovagatran, aplidin, desmopressin, and felypressin listed as potential inhibitors of main proteaseChandel et al. [42]2020In silicoDrug repurposing, molecular dynamics, and dockingFDA-approved drugsNsp9 replicase and spike proteinsConivaptan exhibited the highest binding of the Nsp9 replicase. Tegobuvir exhibited maximum stability along with the highest binding energy at the active site of the spike proteinsChen et al. [43]2020In silicoDrug repurposing, molecular dynamics, and dockingFDA-approved drugsSpike (S)-mediated cell entryCepharanthine, abemaciclib, osimertinib, trimipramine, colforsin, and ingenolChidambaram et al. [44]2020In silicoMolecular dockingCoumarins and their analogsMain proteaseNatural coumarin analog toddacoumaquinone displayed remarkable inhibition ability. Synthetic coumarin analog (1 m) also displayed the comparable inhibition ability main protease in intricate withα-ketoamideChoudhary et al. [45]2020In silicoDrug repurposing, molecular dynamics, and dockingFDA-approved drugsSpike glycoprotein and cellular angiotensin-converting enzyme 2 (ACE2) receptorGR 127935 hydrochloride hydrate, GNF-5, RS504393, TNP, and eptifibatide acetate were found binding to virus binding motifs of ACE2 receptor. KT203, BMS195614, KT185, RS504393, and GSK1838705A were identified to bind at the receptor-binding site on the viral S proteinClemente et al. [46]2021In silicoMolecular docking, molecular dynamic (MD) simulationsIbuprofenMain proteaseRacemic mixtures of the ibuprofen enantiomers might be a potential treatment for main proteaseCosic et al. [47]2021In silicoExtended resonant recognition model (RRM)IvermectinSpike proteinsIvermectin could interfere with activity of spike proteinsDaoud et al. [48]2021In silicoStructure-based pharmacophore approach, molecular docking, and repurposing studiesFDA-approved drugsMain proteaseLopinavir, remdesivir, ritonavir, saquinavir, and raltegravir were successfully docked into the binding site of main proteasede Oliveira et al. [49]2021In silicoMolecular modeling and virtual screening and repurposing studies9091 FDA-approved drugsSpike protein24 best scored ligands (14 traditional herbal isolates and 10 approved drugs) as potential candidates to inhibit the S proteinQuinupristin, nilotinib, acetyldigitoxinDelijewski and Haneczok [50]2021In silicoSupervised machine learning model and repurposing studiesFDA-approved drugsAgainst SARS-CoV-2Zafirlukast as the best repurposing candidate for COVID-19Dey et al. [51]2021In silicoVirtual database screening, molecular docking, all-atom molecular dynamic simulation, and MM-PBSA analysisTretinoin, mefenamic acid, ondansetron, and artemetherEnvelope (E) proteinTretinoin as a potential SARS-CoV-2 E protein ion channel blocker and virus assembly inhibitorDurdagi [52]2020In silicoMolecular modeling approach in virtual drug screening repurposing studyFDA-approved drugsType 2 transmembrane serine protease (TMPRSS2)Benzquercin as strong TMPRSS2 inhibitorDurdagi et al. [53]2020In silicoMolecular docking, MM-GBSA-based predictions, and molecular dynamic repurposing studyFDA-approved drugsMain protease and spike receptor-binding domain bound with ACE2 COVID-19 target proteinsPimelautide, rotigaptide, telinavir, ritonavir, pinokalant, terlakiren, cefotiam, and cefpiramide as SARS-CoV-2 main protease inhibitors. Denopamine, bometolol, naminterol, rotigaptide, and benzquercin as potential ACE2/spike protein domain inhibitorsEleftheriou et al. [54]2020In silicoMolecular docking34 approved and on-trial protease inhibitorsMain proteaseHCV protease, DPP-4,α-thrombin, and coagulation factor Xa known inhibitorsElmezayen et al. [55]2021In silicoMolecular modeling approach in virtual drug screening repurposing studyCommercially available drugs and ZINC15 libraryMain proteasesFour potential inhibitors against Mpro enzyme, two available drugs (Talampicillin and Lurasidone), and two novel drug-like compounds (ZINC000000702323 and ZINC000012481889)Encinar et al. [56]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study9000 US Food and Drug Administration (FDA)-approved investigational and experimental drugs from the DrugBank repositoryS-Adenosyl-L-methionine-binding pocket of nsp16, [2] the unique “activating surface” between nsp16 and nsp10, and [3] the RNA-binding groove of nsp16Tegobuvir, sonidegib, siramesine, antrafenine, bemcentinib, itacitinib, or phthalocyanine antagonism of SARS-CoV-2 RNAs lacking 20-O-methylationFarag et al. [57]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study2000 FDA-approved drugsMain proteaseDarunavir, nelfinavir, and saquinavir bound to the central site of main protease substrate-binding pocket rosuvastatin, montelukast, and the anti-histaminic fexofenadine bound to the terminal site of main protease substrate-binding pocketStarting point for further in vitro and in vivo testingFeng et al. [58]2020In silicoMolecular modeling approach in virtual drug screening and repurposing studyFDA-approved drugsSpike proteinEltrombopag possesses a high binding affinity to S protein plus human ACE2Ferraz et al. [59]2020In silicoLigand and structure-based virtual screening, repurposing studyFDA-approved drugsMain proteaseTwo oral (bedaquiline and glibenclamide) and one buccal drug (miconazole)Fischer et al. [60]2020In silicoMolecular docking approach in virtual drug screening and repurposing studyOver 606 million compoundsMain protease12 purchasable compounds, with binding affinity to the target protease the natural compounds (−)—taxifolin and rhamnetin as potential inhibitors of main proteaseGimeno et al. [61]2020In silicoMolecular modeling approach in virtual drug screening and repurposing studyFDA-approved drugsMain proteasePerampanel, carprofen, celecoxib, alprazolam, trovafloxacin, sarafloxacin, and ethyl biscoumacetate. Carprofen and celecoxibInitiative for in vitro testingGuo et al. [62]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study single-cell RNA sequencingUS FDA-approved drugsAgainst SARS-CoV-2281 FDA-approved drugs that have the potential to be effective against SARS-CoV-2 infection, 16 of which are currently undergoing clinical trials to evaluate their efficacy against COVID-19Including the HIV protease inhibitor lopinavir/ritonavir combination (phase 4), glucocorticoid receptor agonist dexamethasone (phase 3/4), DNA replication inhibitor niclosamide (phase 2/3), antineoplastic agent lenalidomide (phase 4), and calcineurin inhibitor tacrolimus (phase 3), ABT-737 (BCL inhibitor), brefeldin-A (protein synthesis inhibitor), indirubin (CDK inhibitor), TPCA-1 (IKK inhibitor), lopinavir (HIV protease inhibitor), GW-441756 (growth factor receptor inhibitor), treprostinil (prostacyclin analog), tyrphostin-AG-1478 (EGFR inhibitor) and epoxycholesterol (LXR agonist), fostamatinib (SYK inhibitor), VER-155008 (HSP inhibitor), KU-0063794 (MTOR inhibitor), PIK-90 (PI3K inhibitor), linsitinib (IGF-1 inhibitor), TAK-715 (p38 MAPK inhibitor), Y-27632 (Rho-associated kinase inhibitor), AZ-628 (RAF inhibitor), and lestaurtinib (FLT3 inhibitor)Gupta et al. [63]2020In silicoMolecular modeling approach in virtual drug screening and repurposing studyFDA-approved drugsMain proteaseCobicistat is the most efficient inhibitor of Mpro both in silico and in vitroHuynh et al. [64]2021In silicoDocking and molecular dynamics and repurposing studyFDA-approved drugsPapain-like proteaseThe chances of drug repurposing for PLpro might be lowIbrahim et al. [65]2020In silicoMolecular dynamic simulations, molecular docking, MM-GBSA analysis, and repurposing studyDrugBank databaseMain proteaseDB02388 and cobicistat (DB09065)Iftikhar et al. [66]2020In silicoMolecular modeling approach in virtual drug screening and repurposing study4574 compounds also containing FDA-approved drugsRdRp, main protease, and helicaseRimantadine, bagrosin, and grazoprevir showed binding to main protease. Casopitant is a neurokinin-1 receptor that showed binding to RdRp. Meclonazepam and oxiphenisatin showed specific interactions with helicaseJain and Mujwar [67]2020In silicoComputational drug repurposing docking simulations2880 FDA-approved drugsMain proteaseMetocurine, dihydroergotoxine, imatinib, daunorubicin, bromocriptine, irinotecan, azelastine, gestodene, adapalene, and simvastatinMetocurine was chosen as a safe and effective drug candidate for developing therapy against the viral Mpro enzyme of SARS-CoV-2 for the treatment of COVID-19Jarvis et al. [68]2020In silicoTier-based scoring system repurposing studyClinically developed drugsPotential repurposing against COVID-19Four drug classes (antimalarial amino-quinolones, selective estrogen receptor modulators (SERMs), low potency tricyclic antipsychotics, and tricyclic antidepressants) as potential drug candidates for COVID-19The tricyclic antipsychotics and tricyclic antidepressants were further excluded based on a high adverse event profileKadioglu et al. [69]2021In silicoRepositioning/virtual drug screening, molecular docking, and supervised machine learning algorithm drug repositioningFDA-approved drug natural compound dataset ZINC databaseSpike protein, nucleocapsid protein, and 2′-o-ribose methyltransferaseConivaptan, paritaprevir, simeprevir, dihydroergotamine, ZINC000027215482, ZINC000252515584, loniflavone, procyanidinKandeel et al. [70]2020In silicoDrug repurposing molecular dynamic (MD) simulations followed by molecular mechanics/generalized born surface area (MM/GBSA) binding energy calculations1697 clinical FDA-approved drugsPapain-like proteasePhenformin, quercetin, and ritonavirPhenformin was more stable than quercetin and ritonavirKandwal and Fayne [71]2020In silicoRepurposing drug computational design pharmacophore featuresIn-development/approved drugsViral nucleocapsid and nonstructural proteinsIsepamicin and streptomycin (nsp3); coenzyme-I, rutin, epigallocatechin gallate-(-), and procyanidin-b-2 (nsp7/nsp8/nsp12); paromomycin (nsp10/nsp16); olomoucine, sapropterin, tetrahydrofolic acid, INS316, and adenosine phosphate (nsp15); varespladib, hexanoic acid, citric acid, OSI-027, MK-5108, stepronin, calcium gluceptate, CPP, pirenoxine, midafotel, and maltobionic acid (nucleocapsid)Khan et al. [72]2020In silicoDrugs repurposing molecular dynamic simulation31 FDA-approved anti-HIV drugs, and traditional Chinese medicines (TCM) databaseMain proteaseSaquinavir and TCM5280805Khan et al. [73]2020In silicoDrugs repurposing molecular docking23 prospective drug candidatesMain proteaseEpirubicin, vapreotide, and saquinavir exhibited better binding affinitySynergistic interactionKouznetsova et al. [74]2020In silicoDrugs repurposing molecular dockingFDA-approved drugsPapain-like proteaseInhibitors of HIV, hepatitis C, and cytomegalovirus (CMV) demonstrated some activityKrishnaprasad et al. [75]2020In silicoDrugs repurposing molecular dockingFDA-approved library of drugsRNA-dependent RNA polymerasePitavastatin, ridogrel, and rosoxacin displayed superior binding with the active siteKumar et al. [76]2020In silicoDrugs repurposing docking and molecular dynamic (MD) simulations combined with molecular mechanics/generalized born surface area (MM/GBSA)12 FDA-approved drugs (darunavir, indinavir, saquinavir, tipranavir, diosmin, hesperidin, rutin, raltegravir, velpatasvir, ledipasvir, rosuvastatin, and bortezomib)Main proteaseSaquinavir as a potent inhibitor of dimeric main proteaseKumar et al. [77]2020In silicoDrugs repurposing molecular docking molecular dynamic simulations MM/GBSAWithaferin A (Wi-A), withanone (Wi-N) (active withanolides of ashwagandha), and caffeic acid phenethyl ester (CAPE, bioactive ingredient of propolis)Main proteaseWi-N and CAPE possess the potential to inhibit the functional activity of main proteaseKumar et al. [78]2020In silicoDrugs repurposing molecular dockingFDA-approved drugsMain proteaseLopinavir-ritonavir, tipranavir, and raltegravir show the best molecular interaction with the main proteaseKumar et al. [79]2020In silicoDrugs repurposing molecular docking molecular dynamic simulationsFDA-approved library of drugsMain proteaseHyaluronic acid and acarbose show strong interactions with catalytic site residues of main proteaseLi et al. [80]2020In silicoDrug repurposing free energy perturbation-based absolute binding free energy (FEP-ABFE) predictionsVirtual screening of existing drugsMain protease25 drugs were predicted, and 15 were confirmed as potent inhibitors of SARS-CoV-2 main protease. The most potent one is dipyridamole. Hydroxychloroquine (ki = 0.36μM) and chloroquine (ki = 0.56 μM) were also found to potently inhibit main proteaseLiang et al. [81]2021In silicoDrug repurposing molecular docking2,631 FDA-approved small moleculesMultiple main proteins29 drugs that could actively interact with two or more target proteins, with 5 drugs (avapritinib, bictegravir, ziprasidone, capmatinib, and pexidartinib) being common candidates for all four key host proteins and 3 of them possessing the desirable molecular propertiesLokhande et al. [82]2021In silicoDrugs repurposing molecular docking molecular dynamic simulationsFDA-approved drugsMain proteaseMitoxantrone, leucovorin, birinapant, and dynasoreMahanta et al. [83]2020In silicoDrugs repurposing molecular docking molecular dynamic simulationsU.S. Food and Drug Administration-approved antimicrobial drugsMain proteaseViomycinMahdian et al. [84]2021In silicoDrugs repurposing molecular docking molecular dynamic simulationsFDA-approved drugsViral entry receptors (ACE2 and CD147) and integral enzyme of the viral polymerase (RdRp)Ledipasvir, estradiol benzoate, and vancomycin and paritaprevirMarak et al. [85]2020In silicoRepurposing drug homology modeling molecular docking108 FDA-approved antiparasitic and anti-inflammatory drugs10 SARS-CoV-2 targets (PLpro, 3CLpro, RdRp, spike, helicase, NSP1, NSP3, NSP4, NSP9, and NSP16-NSP10)Ivermectin, atovaquone, posaconazole, doxycycline, moxidectin, amphotericin B, chlortetracycline, spiramycin, sulfasalazine, parecoxib, and etoricoxib exhibited good binding affinitiesMohapatra et al. [86]2020In silicoRepurposing drug machine learning (ML) technologyFDA-approved drugsAgainst COVID-1910 FDA-approved commercial drugs that can be used for repurposing amprenavir would probably be the most effective drug based on the selected criteriaMolavi et al. [87]2021In silicoRepurposing drug molecular docking1760 FDA-approved drugsRNA-dependent RNA polymerase (RdRp) and main proteaseNilotinib, imatinib, and dihydroergotamine for 3clpro and dexasone and raltegravir for RdRp. Raltegravir, an anti-HIV drug, was observed to be the best compound against RdRp based on docking binding energy dihydroergotamine is a suitable candidate for main proteaseMulgaonkar et al. [88]2020In silico/in vitroRepurposing drug molecular dockingFDA-approved drugsSpike glycoproteinBCR-ABL tyrosine kinase inhibitor, imatinib, inhibits SARS-CoV-2Via fusion inhibitionMycroft-West et al. [89]2020In silicoRepurposing drug molecular docking molecular dynamic simulationsHeparinSpike (S1) protein receptor-binding domainInhibition of viral infection arises from an overlap between the binding sites of heparin/HS on S1-RBDRepurposing heparin and its derivatives as antiviral agents against SARS-CoV-2Nayarisseri et al. [90]2020In silicoShape-based machine learning assisted by molecular docking and molecular dynamic simulations. ADMET studies31 repurposed compoundsMain proteaseRemdesivir, valrubicin, aprepitant, and fulvestrantThe novel compound nCorv-EMBS herein proposed stands as a promising inhibitor to be evaluated further for COVID-19 treatmentOdhar et al. [91]2020In silicoMolecular docking molecular dynamic simulations1615 FDA-approved drugsMain proteaseConivaptan azelastineOrtega et al. [92]2020In silicoRepurposing drug molecular dockingFamotidineAgainst SARS-CoV2Famotidine could interact within the catalytic site of the three proteases associated with SARS-CoV2 replicationWeak binding affinity could be reached only upon intravenous administrationPandey et al. [93]2021In silicoRepurposing drug molecular docking9 flavonoidsSpike glycoproteinBaicalinParveen and Alnoman [94]2021In silicoMolecular docking molecular dynamic simulation density functional theory (DFT) ADME-ToxFDA-approved anticancer drugs (capmatinib, pemigatinib, selpercatinib, and tucatinib)Spike glycoprotein (S1) and the main proteasePotential of selected anticancer drugs for plausible drug development to fight COVID-19Capmatinib, pemigatinib, selpercatinib, and tucatinibPeele et al. [95]2020In silicoMolecular docking molecular dynamic simulationsUSFDA-approved drugs, plant-derived natural drugsMain proteaseLopinavir, amodiaquine, theaflavin digallatePinzi et al. [96]2021In silicoDrug repurposing molecular docking molecular mechanic Poisson–Boltzmann surface area (MM-PBSA)DrugBank databaseMain protease22 candidates with putative SARS-CoV-2 Mpro inhibitory activity. Enalkiren, ethylsulfonamide-D-Trp-Gln-p-aminobenzamidine, delparantag ritonavir and lopinavir, saquinavirBeneficial polypharmacological effectsPokhrel et al. [97]2020In silicoDrug repurposing molecular dynamic simulationsUS Food and Drug Administration (FDA)-approved drugsRNA-dependent RNA polymeraseQuinupristin is particularly interesting because it is expected to bind across the RNA tunnel, blocking access from both sidesQuinupristin represents a potential anti-SARS-CoV-2 therapeuticRay et al. [98]2020In silicoDrug repurposing intramolecularly quenched fluorescence (IQF) peptide substrate774 FDA-approved drugsMain proteaseEthacrynic acid, naproxen, allopurinol, butenafine hydrochloride, raloxifene hydrochloride, tranylcypromine hydrochloride, saquinavir mesylateSachdeva et al. [99]2020In silicoDrug repurposing molecular dockingAntimalarial drugsSpike protein and main proteaseDoxycycline showed the most effective binding to the spike protein, whereas halofantrine and mefloquine bound effectively with the main proteaseDoxycycline could potentially be a good candidate for repurposing for COVID-19Sang et al. [100]2020In silicoDrug repurposing molecular docking molecular mechanics Poisson–Boltzmann surface area (MM-PBSA)6 approved anti-HIV drugsMain proteaseDarunavirSaxena et al. [101]2021In silico/In vitroDrug repurposing molecular dockingFDA-approved DrugBank databaseSpike proteinErtugliflozin possesses several desired propertiesGood candidate for immediate repurposing for the treatment of COVID-19Setianingsih et al. [102]2020In silicoDrug repurposing molecular docking, molecular dynamic simulations160 potential drugs from therapeutic target database13 protein targets (12 SARS-CoV-2 proteins and 1 human protein)Suramin, the strongest binding affinity against 3 protein targets (spike protein, nucleocapsid protein, ACE2)Suramin is the most potential to bind nucleocapsid and spike protein of SARS-CoV-2Shah et al. [103]2020In silicoDrug repurposing molecular docking61 molecules that are already being used in clinics or under clinical scrutiny as antiviral agentsAgainst the SARS-CoV-237 molecules were found to interact with >2 protein structures of COVID-19. HIV protease inhibitors and RNA-dependent RNA polymerase inhibitors showed promising features of binding to COVID-19 enzymeMethisazone, an inhibitor of protein synthesis; CGP42112A, an angiotensin AT2 receptor agonist; and ABT450, an inhibitor of the nonstructural protein 3-4A, might become convenient treatment option as well against COVID-19Sharma and Mishra [104]2020In silicoDrug repurposing target-based virtual ligand screeningZINC drug database and our own database of natural productsAgainst the SARS-CoV-2Antivirus drugs (ribavirin, valganciclovir, and thymidine), antibacterial drugs (cefpiramide, sulfasalazine, phenethicillin, lymecycline, demeclocycline, doxycycline, oxytetracycline, and tigecycline), anti-asthmatic drugs (montelukast, fenoterol, and reproterol), and hepatoprotective drug silybin have antiviral activity. Natural hesperidin was targeting the binding between spike RBD and human ACE2The natural products, such as flavonoids like neohesperidin, hesperidin, baicalin, kaempferol 3-O-rutinoside, and rutin from different sources, andrographolide, neoandrographolide, and 14-deoxy-11,12-didehydroandrographolide from A. paniculata, and a series of xanthones from the plants of Swertia genus, with antivirus, antibacteria, and anti-inflammation activity could effectively interact with these targets of SARS-CoV-2Shekhar et al. [105]2020In silicoDrug repurposing molecular docking molecular dynamic simulations2,625 FDA-approved small moleculesSpike (S) protein fusion peptide regionChloramphenicol succinate, imipenem, and imidureaSingh et al. [106]2021In silicoDrug repurposing molecular docking molecular dynamic simulations1749 FDA-approved drugsNSP12, a RNA polymerase5 compounds which include 3a (paritaprevir), 3d (glecaprevir), 3h (velpatasvir), 3j (remdesivir), and 3l (ribavirin) had the best binding affinitySinha et al. [107]2021In silicoDrug repurposing systematic pharmacokinetics, drug-likeness, basicity predictions, virtual screening, and molecular dynamic analysisHydroxychloroquine (HCQ), chloroquine (CQ)Spike protein1-[1-(6-Chloroquinolin-4-yl) piperidin-4-yl]piperidin-3-ol and (1r,2R)-2-N-(7-chloroquinolin-4-yl)cyclohexane-1,2-diamine interact with the active site of the spike protein similar to HCQ and CQ, respectively, with augmented safety profileSoni et al. [108]2020In silicoMolecular docking molecular dynamic simulation ADME propertiesRifampicinMain proteaseRifampicin docking score was −7.24 kcal·mol–1, and it can predict as a very good inhibitor of main proteaseTariq et al. [109]2020In silicoDrug repurposing molecular docking molecular dynamic simulations15 antimalarial drugs (including chloroquine) and 2413 US Food and Drug Administration-approved drugsMain protease spike (S) proteinParomomycin with activity against two targets spike protein and protease domainTatar et al. [110]2021In silicoDrug repurposing molecular docking molecular dynamic simulations34 antiviral compoundsRNA-binding domainRapamycin, saracatinib, camostat, trametinib, and nafamostat were the top hit compoundsTejera et al. [111]2020In silicoDrug repurposing quantitative structure-activity relationship (QSAR) mode molecular docking molecular dynamic simulation MM-PBSA methodDrugBank databaseMain proteaseLevothyroxine, amobarbital, and ABP-700Teralı et al. [112]2020In silicoDrug repurposing molecular docking7,173 clinically approved drugAngiotensin-converting enzyme 2 (ACE2)Lividomycin, burixafor, quisinostat, fluprofylline, pemetrexed, spirofylline, edotecarin, diniprofyllineTrezza et al. [113]2020In silicoDrug repurposing docking simulations, with molecular dynamics (MD), supervised MD (SuMD), and steered MD (SMD) simulationsFDA-approved drugsSpike glycoproteinSimeprevir, lumacaftorUgurel et al. [114]2020In silicoDrug repurposing structure-based drug design genome sequences were analyzedFDA-approved drugsHelicase (Nsp13)Cangrelor, fludarabine, folic acid, and polydatin inhibit both the wild-type and mutant SARS-CoV-2 helicaseUnni et al. [115]2020In silicoDrug repurposing molecular docking molecular dynamic simulationsDrugBank and PubChem librarySpike protein (S protein)Bisoxatin (DB09219)A laxative drugVaishali et al. [116]2020In silicoDrug repurposing molecular docking molecular dynamic simulation ADME propertiesFDA-approved compoundsNonstructural protein 9 (Nsp9) replicase and spike proteinsConivaptan exhibited the highest binding energy and maximum stability of the Nsp9 replicase. Tegobuvir exhibited maximum stability along with the highest binding energy at the active site of the spike proteinsVerma et al. [117]2020In silicoDrug repurposing molecular docking molecular dynamic simulation MM-GBSA-based energyFDA-approved drugsMain proteaseTop-ranked drugs including adefovir, lumefantrine, dipyridamole, dihydroergotamine, hexoprenaline, riboflavin (vitamin B2), and pantethine (vitamin B5)Wei et al. [118]2020In silicoDrug repurposing molecular docking molecular dynamic simulationsUS Food and Drug Administration (FDA)-approved drugs from DrugBank and natural compounds from traditional Chinese medicine systems pharmacology (TCMSP)Spike protein (S protein)Digitoxin and bisindigotin in TCMSP had the highest docking scores Forsythiae fructus and Isatidis radix are components of Lianhua Qingwen, and raltegravir had relatively high binding scoresXu et al. [119]2021In silicoDrug repurposing molecular docking molecular dynamic simulationsFDA-approved drugsSpike proteinThymoquinone, a phytochemical compound obtained from the plant Nigella sativa, is a potential drug candidateTable 2
Summary of studies with in silico method that used natural products for novel drug discovery against COVID-19.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAbdelli et al. [120]2021In silicoMolecular dockingIsothymol, thymol, limonene, P-cymene, andγ-terpinene derived from the essential oil of the antiviral and antimicrobial plant Ammoides verticillata (Desf.) Briq.Inhibition of ACE2 cellular receptorIsothymol, a major component of this plant, gives the best docking scores, as good ACE2 inhibitorAbouelela et al. [121]2021In silicoMolecular docking, dynamic simulation, and binding free energy calculationAloeMain protease and spike protein132, 134, and 159 were the best scoring compounds against main protease, while compounds 115, 120, and 131 were the best scoring ones against spike glycoprotein. Compounds 120 and 131 were able to achieve significant stability and binding free energies during molecular dynamic simulationAdem et al. [122]2020In silicoMolecular dockingMedicinal plant-based bioactive compounds (80 flavonoid compounds)Main proteaseHesperidin, rutin, diosmin, apiin, diacetylcurcumin, (E)-1-(2-hydroxy-4-methoxyphenyl)-3-[3-[(E)-3-(2-hydroxy-4-methoxyphenyl)-3-oxoprop-1-enyl]phenyl]prop-2-en-1-one, and beta, beta′-(4-methoxy-1,3-phenylene)bis(2′-hydroxy-4′,6′-dimethoxyacrylophenone have been found as more effective on COVID-19 than nelfinavirAllam et al. [123]2020In silicoMolecular docking, 3D shape similarity study (rapid overlay chemical similarity-ROCS) to the clinically used drugs in COVID-19 patients3′-Hydroxy-4′-methoxy-chroman-7-O-β-d-glucopyranoside 4, ferulic acid heptyl ester 1, naringenin 2, and 4,2′,4′-trihydroxy-6′-methoxychalcone-4′-O-β-d-glucopyranoside 3, which were isolated from peach (Prunus persica (L.) Batsch) fruitsMain protease, spike proteinNaringenin 2 and 4,2′,4′-trihydroxy-6′-methoxychalcone-4′-O-β-d-glucopyranoside 3 have a strong binding mode to a protease receptor and spike protein and also block the inflammatory stormRecommendation of peach fruits in controlling and managing COVID-19 casesAl‐Sehemi et al. [124]2020In silicoMolecular docking31000 natural compounds of the natural product activity and species source (NPASS) librarySpike glycoproteinCastanospermine and karuquinone B were shown to be the best-in-class derivatives in silico able to target an essential structure of the virus and to act in the early stage of infectionAttia et al. [125]2021In silicoMolecular docking10 phenolic antiviralAgainst SARS-CoV-2Hesperidin showed the highest docking scoreHesperidin and its mediated ZnO nanoparticles are willing antiviral agentsAzim et al. [126]2020In silicoVirtual screening methods molecular docking27 plant metabolitesMain protease proteins (MPP), Nsp9 RNA-binding protein, spike receptor-binding domain, spike ectodomain, and HR2 domainAsiatic acid, avicularin, guajaverin, and withaferin showed a maximum binding affinity with all key proteins in terms of lowest global binding energyBhowmik et al. [39]2021In silicoRepurposing drugs, docking, and molecular dynamic simulationOrientin (phytochemical)Inhibitor of SARS-CoV-2 spike and host cell receptor GRP78 bindingBinding of orientin in the overlapping residues of GRP78 binding region of SARS-CoV-2 spike modelAs a promising precautionary or therapeutic measure for COVID-19Çakır et al. [127]2021In silicoMolecular dockingPeptides derived from beta-lactoglobulinInhibit the host cell membrane receptorsAla-Leu-Pro-Met-His-Ile-Arg (ALMPHIR) and Ile-Pro-Ala-Val-Phe-Lys (IPAVFK) peptidesβ-Lactoglobulin (BLG) is the major whey protein of cow and sheep’s milk (∼3 g/l)Chatterjee et al. [128]2021In silicoMolecular dockingHesperidin, kaempferol, quercetin, epigallocatechinPLpro (papain-like protease), RdRp (RNA-dependent RNA polymerase), Mpro or 3cl protease, and spike proteinHesperidin, kaempferol, quercetin, epigallocatechinLead to conclusive data for the treatment of polyphenols, flavonoids, and bioflavonoids against SARS-CoV-2Chikhale et al. [129]2020In silicoMolecular docking, dynamicsPlant Withania somnifera (Indian ginseng)NSP15 endoribonuclease and receptor-binding domain of prefusion spike proteinWithanoside X and quercetin glucoside from W. somnifera have favorable interactions at the binding site of selected proteinsImmunomodulatory, antioxidant, and anti-inflammatory rolesChikhale et al. [130]2021In silicoMolecular docking, dynamics, and network pharmacology analysisSaikosaponinsAdjuvant therapy in the treatment of COVID-19Saikosaponins interact with the proteins CAT gene CAT (catalase) and checkpoint kinase 1 (CHEK1)Possible improvement in immune response towards COVID-19Chikhalet al. [131]2020In silicoMolecular docking, dynamicsAsparagus racemosus (Willd.)NSP15 endoribonuclease and spike receptor-binding domainAsparoside -C and Asparoside -F have good binding with target proteinsAsparagus racemosus holds promise as SARS-CoV-2 (S) and (N) protein inhibitorChowdhury [132]2020In silicoMolecular docking, dynamicsTinospora cordifolia (Giloy)Main proteaseBerberine can regulate main protease protein’s functionDahab et al. [133]2020In silicoMolecular docking10 phenolic compounds of different classes (phenolic acids, flavonoids, and coumarins)Main protease and RNA polymeraseThe top 7 hits are flacourticin [3], sagerinic acid [16], hordatine a [23], hordatine B [134], N-feruloyl tyramine dimer [135], bisavenanthramides B-5 [27], and vulnibactins [38] and have better binding scores than remdesivir, the native ligand in RNA polymerase target (PDB ID: 7bV2)Hordatines are phenolic compounds present in barley and were found to exhibit the highest binding affinity to both protease and polymeraseDas et al. [136]2020In silicoMolecular dockingFlavonoid-based phytochemicals of Calendula officinalisMain proteaseRutin, isorhamnetin-3-O-β-D, calendoflaside, narcissin, calendulaglycoside B, calenduloside, and calendoflavoside have better binding energy than the native ligandRutin and caledoflaside showed better stability, compactness, and flexibilityDebnath et al. [137]2020In silicoSequential E-pharmacophore and structure-based virtual screening (VS)113687 number of commercially available natural compoundsADP-ribose phosphatase6 potential inhibitors having good binding affinity towards active sitesCommercially availableDev and Kaur [138]2020In silicoMolecular dockingEucalyptus essential oilMain proteaseJensenone may represent potential treatment potential to act as main protease inhibitorDuru et al. [139]2021In silicoMolecular dockingOil of Nigella sativa seedReplicase polyprotein 1a, RNA-binding protein of NSP9, ADP ribose phosphatase of NSP3, 3-chymotrypsin-like protease 3CLpro, and RNA-dependent RNA polymerase RdRp, and ACE2-angiotensin-converting enzyme from the Homo sapiensThe binding affinity of caryophyllene oxide was the highest on NSP9 and RdRp targets, whileα-bergamotene gave the best binding affinity on RPIA target. The binding affinity of β-bisabolene on the ACE2 was almost the same as remdesivirEl-Demerdash et al. [140]2021In silicoMolecular dynamic simulations, molecular docking15 guanidine alkaloidsMain protease (Mpro) (PDB ID: 6lu7), spike glycoprotein (PDB ID: 6VYB), nucleocapsid phosphoprotein (PDB ID: 6VYO), membrane glycoprotein (PDB ID: 6M17), and a nonstructural protein (nsp10) (PDB ID: 6W4H)Crambescidin 786 [5] and crambescidin 826 had the highest binding affinities. The examined 15 alkaloids especially 5 and 13 showed promising docking, ADMET, toxicity, and MD resultsElekofehinti et al. [141]2020In silicoMolecular docking studies, molecular dynamics, and ADME/Tox50,000 natural compounds retrieved from IBS databasePapain-like proteaseSTOCK1N-69160 [(S)-2-((R)-4-((R)-2-amino-3-methylbutanamido)-3-(4-chlorophenyl) butanamido)propanoic acid hydrochloride] has been proposed as a novel inhibitor against COVID-19 PLproEl‐Hawary et al. [142]2021In silicoMolecular docking (a combination of metabolomics and in silico approaches)A. terreus, the endophytic fungus associated with soybean rootsMain proteaseAspergillide B1 and 3α-hydroxy-3,5-dihydromonacolin L were found to be potent anti-COVID-19 drug candidatesEmirik [143]2020In silicoMolecular docking, MM-GBSA-based predictions, and molecular dynamicsTurmeric contentsSARS-CoV-2 vital proteinsTurmeric spice has the potential to inhibit the SARS-CoV-2 vital proteins and can be used a therapeutic or protective agent against SARS-CoV-2 via inhibiting key protein of the SARS-CoV-2. Compounds 4, 23, and 6 are the most prominent inhibitor for the main protease, the spike glycoprotein, and RNA polymerase of virus, respectivelyFakhar et al. [144]2020In silicoStructure-based pharmacophore modeling, virtual screening-based PHASE screen score, molecular modelingAnthocyanin derivativesMain protease6 best anthocyanin-derived natural compounds, which could be used as promising lead compounds against main protease SARS-CoV-2Falade et al. [145]2021In silicoMolecular dockingSaponins and tanninsMain proteaseEllagic acid, arjunic acid, theasapogenol B, and euscaphic acid as potential inhibitors of SARS-CoV-2 (Mpro) with better pharmacokinetics and bioavailability compared with remdesivirFitriani et al. [146]2020In silicoMolecular dockingPhytochemical compounds (Moringa oleifera, Allium cepa, Cocos nucifera, Psidium guajava, and Eucalyptus globulus)Main proteaseOleanolic acid in Allium cepa,α-tocotrienol in Cocos nucifera, asiatic acid in Psidium guajava, and culinoside in Eucalyptus globulus were the most recommended compound in each medicinal plantOleanolic acid in Allium cepa found as a potential inhibitor of COVID-19 MproGangadevi et al. [147]2021In silicoMolecular dynamic simulations, molecular dockingLibrary of natural compoundsHost ACE2 receptor with spike RBD domain of SARS-CoV-2Kobophenol A, identified through docking studies, is the first compound that inhibits SARS-CoV-2 binding to cells through blocking S1-RBD to the host ACE2 receptor and thus may serve as a good lead compound against COVID-19Gangarapu et al. [148]2020In silicoMolecular docking online pkCSM and SwissADME Web serverPhytoconstituents of Siddha official formulation kabasura kudineer and novel herbal preparation—JACOMSpike protein37 compounds were screened, and of these, 9 compounds showed high binding affinity against spike proteinSNACK-V formulations could be used for effective treatment of COVID-19Ghosh et al. [149]2020In silicoMolecular dynamic simulations, molecular docking8 polyphenols from green teaMain protease3 polyphenols (epigallocatechin gallate, epicatechin gallate, and gallocatechin-3-gallate) interact strongly with one or both catalytic residues (His41 and Cys145) of main proteaseGhosh et al. [150]2021In silicoMolecular dynamic simulations, molecular docking, MM-GBSA analysisJusticia adhatoda alkaloidsMain protease1 alkaloid (anisotine) had interaction with both the catalytic residues (His41 and Cys145) of Mpro and exhibited good binding affinity (−7.9 kcal/mol)More potent Mpro inhibitor than the two previously recommended antiviral drugs (lopinavir and darunavir)Gorla et al. [151]2020In silicoMolecular dockingEssential flavonoidsSARS-CoV-2 spike glycoprotein receptor-binding domain (RBD-S) and host angiotensin-converting enzyme-2 protease domain (PD-ACE2)Biochanin A and silymarin bind significantly at the active sites of RBD-Sand PD-ACE2Gurung et al. [152]2020In silicoVirtual screening, molecular dockingAntiviral compounds from plantsMain proteaseBonducellpin D was identified as the best lead molecule, which shows higher binding affinityGyebi et al. [153]2021In silicoMolecular docking, ADME/Tox, and Lipinski filter analysisAfrican plants derived alkaloids and terpenoidsMain protease4 nontoxic, druggable plant-derived alkaloids (10-hydroxyusambarensine and cryptoquindoline) and terpenoids (6-oxoisoiguesterin and 22-hydroxyhopan-3-one)Elwakil et al. [154]2021In silicoGas chromatography/mass spectrometry analysis molecular dockingEgyptian propolisRNA-dependent RNA polymerase, spike protein S1, and main proteaseOctatriacontyl pentafluoropropionate is well oriented inside the enzyme pockets, in addition to an excellent binding manner with the active site of the target macromoleculesMenoufia propolis could be a promising candidate in the combat against the pandemic COVID-19Hasan et al. [155]2020In silicoMolecular dockingCompounds present in the plant Solanum surattenseMain protease13 phytochemicals were studied, eight showed very strong binding affinities to main protease, and four showed moderate to strong binding affinitiesHashem [156]2020In silicoMolecular dockingHoneybee and propolisMain protease6 main compounds possess high binding energy with the receptor active site of the main proteaseIbrahim et al. [157]2020In silicoMolecular dynamic simulations, molecular docking, MM-GBSA analysisMolPort database that contains over 100,000 natural productsMain protease9 potent natural products with binding affinities (ΔG binding) >−48.0 kcal/mol four bis([1, 3]dioxolo)pyran-5-carboxamide derivatives were identified as potential drug candidatesMolPort-004-849-765, MolPort-000-708-794, MolPort-002-513-915 and MolPort-000-702-646 are bis([1,3]dioxolo)pyran-5-carboxamide derivativesIbrahim et al. [158]2020In silicoMolecular dynamic simulations, molecular dockingMetabolites present in several common spicesMain proteaseHigh potency of salvianolic acid A and curcumin as main protease inhibitorsSalvianolic acid A as an in silico natural product inhibitor against the SARS-CoV-2 main proteaseIsa et al. [159]2020In silicoDocking and molecular dynamic (MD) simulationExtracts of Zingiber officinale and Anacardium occidentaleMain proteaseSix compounds had good binding energies. CID_9910474 and CID_10503282 had a better stability when compared to other selected phytochemicalsIstifli et al. [160]2020In silicoMolecular dynamics and molecular mechanic Poisson–Boltzmann surface area (MM/PBSA) methods23 phytochemicals belonging to different flavonoid subgroupsSpike glycoprotein cellular proteases (transmembrane serine protease 2 (TMPRSS2), cathepsin B and L (CatB/L)).(−)-Epicatechin gallate interacted strongly with all the proteins studiedEpicatechin gallate can be evaluated as a candidate molecule in drug development studies against 2019-nCoV since it was not the substrate of P-gp (P-glycoprotein), did not inhibit any of the cytochrome Ps, and did not show AMES toxicity or hepatotoxicity on eukaryotic cellsJan et al. [161]2021In silicoCell-based infection assay molecular modeling2,855 small molecules and 190 traditional herbal medicinesMain protease RNA-dependent RNA polymeraseMefloquine, nelfinavir, and extracts of Ganoderma lucidum (RF3), Perilla frutescens, Mentha haplocalyxJo et al. [162]2020In silicoDockingFlavonoidsMain proteaseBaicalin, herbacetin, and pectolinarin have been discovered to block the proteolytic activity. Baicalin showed an effective inhibitory activity against main proteaseJoshi et al. [163]2021In silicoDocking7100 moleculesMain proteaseSeveral natural molecules such asδ-viniferin, myricitrin, taiwanhomoflavone A, lactucopicrin 15-oxalate, nympholide A, afzelin, biorobin, hesperidin, and phyllaemblicin B strongly binds to main proteaseThese molecules also showed strong binding with other potential targets of SARS-CoV-2 infection such as viral receptor human angiotensin-converting enzyme 2 (hACE2) and RNA-dependent RNA polymerase (RdRp)Junior et al. [164]2021In silicoDocking and molecular dynamic (MD) simulationLapachol(1,4-naphthoquinone)SARS-CoV-2 nonstructural proteins (nsps)Lapachol derivatives VI and IX demonstrated the strongest bindingLapachol derivatives are potential ligands for SARS-CoV-2 Nsp9Kar et al. [165]2020In silicoMolecular docking molecular dynamic simulations and analysis of MM-PBSA energyIndian plants including Justicia adhatoda, Ocimum sanctum, and Swertia chirataSpike protein, main protease enzyme Mpro, and RNA-dependent RNA polymerase (RdRp)Anisotine against SARS-CoV-2 spike and Mpro proteins and amarogentin against RdRp as potential inhibitorsKhalifa et al. [166]2020In silicoMolecular docking modeling structural-relationship activity10 anthocyaninsMain proteasePhacelianin, gentiodelphin, cyanodelphin, tecophilinLeading molecules for further optimization and drug development process to combat COVID-19Khalifa et al. [167]2020In silicoMolecular operating environment molecular docking19 structural different hydrolysable tanninsMain proteasePedunculagin, tercatain, and castalinKhan et al. [168]2020In silicoMolecular dockingMarine natural productsMain proteaseC-1 (PubChem CID 11170714) exhibited good activityKiran Raj et al. [169]2020In silicoMolecular dockingC-Phycocyanin of Spirulina platensisNonstructural proteins 12C-Phycocyanin inhibits the active site of nsp12Krupanidhi et al. [170]2020In silicoMolecular docking molecular dynamic simulation ADME along with toxicity analysisPhytochemical constituents of Tinospora cordifoliaMain proteaseTinospononeKumar et al. [171]2020In silicoMolecular docking molecular dynamic simulations and analysis of MM-PBSA energyNovel natural metabolitesMain proteaseUrsolic acid, carvacrol, and oleanolic acid couldKumar et al. [172]2021In silicoMolecular docking, ADMET, and molecular dynamic simulationsPhytoconstituents from natural herbsMain proteaseLaurolitsineLaurolitsine, an active constituent of roots of Lindera aggregataKumar et al. [173]2021In silicoMolecular docking molecular dynamic simulations and analysis of MM-PBSA energyStrychnos nux-vomicaMain proteaseDemethoxyguiaflavine, strychnoflavineLi et al. [174]2021In silicoNetwork pharmacology and in vitro experiment verification molecular dockingMaxing Shigan decoction (MXSGD)ACE2, Mpro, and RdRpThe components with strong potential affinity (top 10) with ACE2, Mpro, and RdRp are mainly from Glycyrrhiza uralensis (Chinese name: Gancao) and Semen armeniacae amarum (Chinese name: Kuxingren). Among them, amygdalin was selected as the optimal candidate component binds to all three key targets, and euchrenone, glycyrrhizin, and glycyrol also exhibited superior affinity interactions with ACE2, Mpro, and RdRp, respectivelyMulticomponent, multitarget, and multi-approach interventionMaiti and Banerjee [175]2021In silicoBioinformatic molecular dockingTea flavonoid catechin productsSpike glycoproteinsEpigallocatechin gallate and theaflavin gallate interact better than hydroxychloroquineMahmud et al. [176]2021In silicoMolecular docking molecular dynamic simulation MM-GBSA scores3063 compounds from more than 200 plants from the Asian regionMain proteaseCurcumin, gartanin, robinetinMahmud et al. [177]2020In silicoMolecular docking molecular dynamic simulation MM-GBSA scoresPlant-derived natural compoundsMain proteaseCarinol, albanin, myricetinMesli et al. [178]2021In silicoMolecular docking molecular dynamic simulationsLeaves of Corchorus olitorius Linn.Angiotensin-converting enzyme 2Méthyl-1,4,5-tri-O-caféoyl quinate has a stronger bond, high affinity, and gives the best docking scores compared with the co-crystallized inhibitor (PRD_002214) of the enzyme ACE2Mohammadi et al. [179]2020In silicoMolecular dockingMarine polypeptides were isolated from the hydrolysate of Pacific oysterMain proteaseThe score of Leu-Leu-Glu-Tyr-Ser-Ileu ligand was higher than remdesivirPacific oyster compounds may have the potency to be evolved as an anti-COVID-19 main proteaseMurugan et al. [180]2020In silicoMolecular docking molecular dynamic simulation MM-GBSA scoresAndrographis paniculata phytochemicals3 nonstructural proteins (3 L main protease (3CLpro), papain-like proteinase (PLpro) and RNA-directed RNA polymerase (RdRp)), and a structural protein (spike protein (S))Neoandrographolide (AGP3) has shown promising binding affinity towards all the four targetsNaik et al. [181]2020In silicoMolecular docking molecular dynamic simulation ADME propertiesNatural product activity and species source (NPASS) databaseEndoribonuclease exoribonuclease RNA-dependent RNA polymerase (RdRp) methyltransferase and main protease21 compounds showed maximum docking scores NPC214620, NPC52382, and NPC270578 are targeting five, four, and three-drug targets, respectivelyMultitarget-based drug designNarkhede et al. [182]2020In silicoMolecular docking molecular dynamic simulationsNatural productsMain proteaseGlycyrrhizin, bicyclogermacrene, tryptanthrin,β-sitosterol, indirubin, indican, indigo, hesperetin, chrysophanic acid, rhein, berberine, and β-caryophylleneInteractions with the COVID-19 main protease were highest in the case of glycyrrhizin and rheinNivetha et al. [183]2020In silicoMolecular docking molecular dynamic simulation MM-PBSASeselin purified from the leaf extracts of Aegle marmelosSpike protein S2, main protease, and free enzyme of the SARS-CoV-2Seselin had inhibitory potential over multiple SARS-CoV-2 targetsOgunyemi et al. [184]2020In silicoMolecular docking molecular dynamic simulation ADME properties226 bioactive compounds from African herbs and medicinal plantsRNA-dependent RNA polymeraseDrugable alkaloids (10′-hydroxyusambarensine, cryptospirolepine, strychnopentamine) and flavonoids (usararotenoid A and 12α-epi-millettosin)Padhi et al. [185]2021In silicoMolecular docking ADME properties415 natural metabolites isolated from several plants, fungi, and bacteriaSpike glycoprotein (S1) and the main proteaseFusaric acid, jasmonic acid, jasmonic acid methyl ester, putaminoxin, putaminoxins B and D, and stagonolide K were predicted to have considerable (ADME) and safety indicesJasmonic acid and putaminoxins B and D bind best to main proteasePandey et al. [186]2020In silicoMolecular docking molecular dynamic simulation ADME properties10 potential naturally occurring compounds (flavonoids/non-flavonoids)Spike glycoproteinFisetin, quercetin, and kaempferol consist of drug-likeness propertyKumar et al. [187]2020In silicoMolecular dockingKabasura kudineer and thonthasura kudineer are two Siddha formulationsSpike glycoproteinCucurbitacin B (−112.09), cardiofolioside (−111.5), apigenin (−98.84), and pyrethrin (−92.98) were observed as more effective with less bind energiesKabasura kudineer could be a potential Siddha medicine for COVID-19Rahman et al. [188]2021In silicoMolecular docking ADMET propertiesRutinMain protease (Mpro), RNA-dependent RNA polymerase (RdRp), papain-like protease (PLpro), and spike (S) proteinSignificant binding of rutin with Mpro, RdRp, PLpro, and S proteins. Main protease exhibited the strongest binding affinityOptimal solubility, nontoxic, and noncarcinogenic propertiesRahman et al. [189]2020In silicoMolecular operating environment (MOE) ligand-based pharmacophore approach and a molecular docking-based screeningNatural product activity and species source (NPASS)Type II transmembrane serine protease (TMPRSS2)85 compounds with molecular docking comparable to or greater than that of the standard inhibitor (camostat mesylate) were identified. The top 12 compounds with the most favorable structural features were studied. The low-molecular-weight compound NPC306344 showed significant interaction with the active site residues of TMPRSS2Rakib et al. [190]2020In silicoMolecular dockingBioactive phytocompounds isolated from Tinospora crispaMain proteaseThe top nine hits might serve as potential anti-SARS-CoV-2 lead molecules, with three of them exerting biological activityRamadhan et al. [191]2020In silicoMolecular dockingEtlingera elatior plantMain proteaseErgosterol peroxide sitostenoneRangsinth et al. [192]2021In silicoMolecular docking ADMET propertiesNatural products isolated from edible and medicinal mushroomsMain protease6 of 25 compounds are the best drug-like property candidates, including colossolactone VIII, colossolactone E, colossolactone G, ergosterol, heliantriol F, and velutinRivero-Segura et al. [193]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesMexican natural productsAgainst the SARS-CoV-2Quercetin, riolozatrione, and cichoriin target the key proteins of SARS-CoV-2Cichoriin reaches higher lung levels (100 mg/kg, IV); therefore, it may be considered in developing therapeutic toolsSelvaraj et al. [194]2020In silicoHomology modeling and molecular dynamic (MD) simulation MM/GBSA, MD simulations, and PCA calculationsTraditional Chinese medicine (TCM) databaseNsp 14 guanine-N7 methyltransferase (N7-MTase)TCM 57025, TCM 3495, TCM 5376, TCM 20111, and TCM 31007 are the compounds from the TCM database, which can occupy and interact nicely with the substrate-binding site of N7-MTaseSharma and Kaur [195]2021In silicoMolecular docking, protein interaction calculator ADME studies12 bioactive molecules present in essential oils of eucalyptus plant leavesSpike (S) proteinToruatoneSharma [196]2020In silicoMolecular docking, protein interaction calculatorEucalyptol (1,8 cineole), an essential oil component from eucalyptus oilMain proteaseEucalyptol may represent potential treatment potential to act as main protease inhibitorEffective binding of eucalyptol to COVID-19 proteinaseSharma and Kaur [197]2020In silicoMolecular docking, protein interaction calculatorJensenone, an essential oil component from eucalyptus oilMain proteaseJensenone may represent potential treatment potential to act as main protease inhibitorShawan et al. [198]2021In silicoPharmacophore study molecular docking molecular dynamic simulation ADME properties43 flavonoids of 7 different classesAgainst the SARS-CoV-2Luteolin and abyssinone II were found to develop successfully docked complex within the binding sites of target proteinsSindhu et al. [199]2020In silicoMolecular dockingClerodendrum paniculatum leavesMain proteaseClerodolSingh et al. [200]2021In silicoMolecular docking and structural dynamic studiesTea (Camellia sinensis) polyphenolsNonstructural protein 16 (NSP16)Theaflavin compound demonstrated lower binding free energy in comparison with the standard molecule sinefunginSingh et al. [201]2020In silicoMolecular docking and structural dynamic studies molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) ADME propertiesPolyphenolsRNA‐dependent RNA polymerase (RdRp)EGCG, theaflavin (TF1), theaflavin-3′-O-gallate (TF2a), theaflavin-3′-gallate (TF2b), theaflavin 3,3′-digallate (TF3), hesperidin, quercetagetin, and myricetin strongly bind to the active site of RdRpEGCG, TF2a, TF2b, and TF3 can inhibit RdRp and represent an effective therapy for COVID-19Srimathi et al. [202]2020In silicoMolecular dockingTraditional herbal medicine: apo-quinine, catechin, cinchonidine, cinchonine, cupreidine, epicatechin, epiprocurcumenol, epiquinine, procurcumenol, quinidine, quinine, zedoaronediol, procurcumadiolAgainst the SARS-CoV-2Epicatechin, apo-quineSubbaiyan et al. [203]2020In silicoMolecular dockingActive constituents present in common herbsSpike (S) proteinEpigallocatechin gallate (EGCG) was found to have the highest binding affinity with the viral S protein, followed by compounds, “F” (curcumin), “D” (apigenin), and “E” (chrysophanol)Green teaSurti et al. [204]2020In silicoMolecular docking molecular dynamic simulationsIlimaquinone (marine sponge metabolite)Spike receptor-binding domain, RNA-dependent RNA polymerase, Nsp10, Nsp13, Nsp14, Nsp15, Nsp16, main protease, and papain-like proteaseIlimaquinone exhibited promising inhibitory potential against all the SARS-CoV-2 target proteins, as evident from the binding energiesMost promising inhibitory candidate against the SARS-CoV-2 papain-like proteaseTao et al. [205]2020In silicoNetwork pharmacology and molecular docking.Huashi Baidu formula (HSBDF): ChineseAgainst the SARS-CoV-2Baicalein and quercetin were the top two compounds of HSBDF, which had high affinity with ACE2Regulate multiple signaling pathways through ACE2Umar et al. [206]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesAzadirachta indica, Mangifera indica, and Moringa oleifera: African plantsMain proteaseMost of the active phytocomponents of the study plants exhibited relative inhibitory potentials against main protease and preferred pharmacological features when compared with hydroxychloroquineCaffeic acid, chlorogenic acid, catechin, ellagic acid, gallic acid, etc.Umesh et al. [207]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesChemical compounds from Indian spicesMain proteaseCarnosol exhibited the highest binding affinity for arjunglucoside-I and rosmanol showed a strong and stable binding affinity with favorable ADME propertiesYang et al. [208]In silico2020High-throughput virtual screeningNatural Products Research Laboratories (NPRL)Main proteaseCurcuminoid derivatives (including NPRL334, NPRL339, NPRL342, NPRL346, NPRL407, NPRL415, NPRL420, NPRL472, and NPRL473) display strong binding affinity to COVID-19 3Lpro polyproteinNPRL-334 revealed the strongest binding affinityYu et al. [209]2020In silicoMetascape analysis protein docking molecular dockingMongolian medicineSARS-CoV-2 S protein RBD domain253 active components were predicted. Phillyrin and chlorogenic acid can effectively prevent the combination of SARS-CoV-2 S protein and ACE2 at the molecular levelZhang et al. [210]2020In silicoMolecular docking molecular dynamic simulation ADME property network pharmacology analysisChinese herbal medicinesAnti-2019-nCoV activity13 compounds that exist in traditional Chinese medicines were found to have potential anti-2019-nCoV activity. 125 Chinese herbs were found to contain 2 or more of these 13 compounds. Of these 125 herbs, 26 are classically cataloged as treating viral respiratory infectionsRegulating viral infection, immune/inflammation reactions, and hypoxia responseTable 3
Summary of other studies with in silico method.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAbu-Melha et al. [211]2020In silicoMolecular docking combined with molecular dynamic simulation (MDS)Hydrazones, pyrazoles, and pyrazines bearing thiazole moietyMain proteaseThe average binding affinities of the compounds 3a, 3b, and 3c (−8.1 ± 0.33 kcal/mol, −8.0 ± 0.35 kcal/mol, and −8.2 ± 0.21 kcal/mol, respectively) are better than that of the positive control nelfinavirAghaee et al. [212]2021In silicoPharmacophore model molecular docking combined with molecular dynamic simulation (MDS), MM/PBSA, ADME studiesPharmit websiteMain proteaseML188, nelfinavir, lopinavir, ritonavir, andα-ketoamideAhmad et al. [213]2021In silicoStructure-based virtual screening (SBVS) of ASINEX antiviral library, molecular dynamic (MD) simulationsASINEX antiviral libraryMain proteaseSCHEMBL12616233, SCHEMBL18616095, and SCHEMBL20148701 compounds conformation with main protease show good stability after initial within active cavity moves, a rich intermolecular network of chemical interactions, and reliable relative and absolute binding free energiesBBB_26580140 lead and its similar analogs to be explored in vivo lead moleculesAhmed et al. [214]2020In silicoMolecular docking, molecular dynamics, and structure-activity relationship76 prescription antiviral drugsRNA-dependent RNA polymerase (RdRp) and main protease (Mpro)Raltegravir, simeprevir, cobicistat, and daclatasvir have higher binding energy and strong interaction with active sites of the receptor proteins (with a precision of 85%)Alabboud and Javadmanesh [215]2020In silicoMolecular docking combined with molecular dynamic simulation88 conventional drugs, 16 vitamins, and 63 natural (plant)Main proteaseVarious vitamins (B9, A, K, and E vitamins) exhibited a significantly strong interaction with the studied receptor. Pleconaril, adefovir dipivoxil, and stavudine in addition to plant-based compounds such as curcumin (Curcuma longa), anolignan A (Anogeissus acuminata), and phyllamyricin B (Phyllanthus myrtifolius) had strong ligand-protein interactionsAlamri et al. [216]2020In silicoStructure-based virtual screening coupled with all-atom molecular dynamic (MD) simulationsProtease inhibitors database composed of ∼7,000 compoundsPapain-like proteaseADM_13083841, LMG_15521745, and SYN_15517940 showed stable conformation and interacted well with the active residues of papain-like proteaseAlexpandi et al. [217]2020In silicoMolecular docking113 quinoline drugsMain protease, RNA-dependent RNA polymerase (RdRp) inhibitors spike-RBD-ACE2 inhibitorElvitegravir and oxolinic acid are able to interact with the NTP entry channel and thus interfere with the RNA-directed 5′–3′ polymerase activity of SARS-CoV-2 RdRp. Rilapladib is the only quinoline that can interrupt the spike-RBD-ACE2 complexQuinoline, 1,2,3,4-tetrahydro-1-[(2-phenylcyclopropyl)sulfonyl]-trans-(8CI), saquinavir, elvitegravir, oxolinic acid, and rilapladib are suggested for the treatment of COVID-19Al‐Sehemi et al. [218]2020In silicoMolecular docking, MD simulation, and molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) resultsPhenyl furoxan, an exogenous nitric oxide donorMain proteaseSpiro-isoquinolino-piperidine-furoxan moieties can be used as an effective ligand for main protease inhibition due to the presence of key isoquinolino-piperidine skeleton with additional NO effectAl-Shar’i [219]2020In silicoMolecular docking, MD simulation, and molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) resultsDifferent databasesMain protease9 compounds with different chemotypesBadavath et al. [220]2020In silico/in vitroMolecular docking, molecular dynamics, and structure-activity relationship studiesScreening of 118 compounds with 16 distinct heterocyclic moieties in comparison with 5 natural products and 7 repurposed drugsMain proteaseOxidiazoles (A2 and A4) derivatives have the best docking scores. Structure-activity relationship studies showed a good comparison with a known active main protease and repurposed drug ebselen with an IC50 value of −0.67 μMBarros et al. [221]2020In silicoMolecular docking24 ligandsSARS-CoV-2 receptors, Nsp9 replicase, main protease (Mpro), NSP15 endoribonuclease, and spike protein (S protein) interacting with human ACE2Antimalarial drug metaquine and anti-HIV antiretroviral saquinavir interacted with all the studied receptorsBasit et al. [222]2020In silicoProtein-protein docking and molecular dynamic simulationTruncated version of human ACE2 (tACE2)S GlycoproteintACE2 provides a high-affinity protein inhibitor for S glycoproteinBattisti et al. [223]2020In silicoPharmacophore‐based screening, docking consensus approach (DCA), molecular dynamic simulations, common hit approach (CHA)Aldrich Market Select (AMS) database from ChemNavigator/Sigma‐Aldrich with over 8 million unique chemical structuresAgainst SARS-CoV-210 compounds with high coronavirus inhibition potentialLead moleculesBenítez-Cardoza and Vique-Sánchez [224]2020In silicoMolecular docking500,000 compoundsPotential inhibitors of the interaction between ACE2 and SARS-CoV-2 (RBD)20 compounds were determined by docking focused on the region of interaction between ACE2 and RBDCava et al. [225]2020In silicoGene Ontology and enrichment analysis protein-protein interaction (PPI) network virtual screening method—Against SARS-CoV-2A protein-protein interaction network of 193 genes, 22 interactions, and 36 potential drugs for future treatment strategies including nimesulide, fluticasone propionate, thiabendazole, photofrin, and didanosineOnly didanosine is a real antiviral drug, while the others are mostly anti-inflammatoryChen et al. [226]2020In silicoCrystal structure, virtual screening7173 purchasable drugs (drugs-lib), with 4574 unique compounds and their stereoisomersMain protease16 candidates for consideration, ledipasvir velpatasvirChoudhary et al. [227]2020In silicoMolecular docking, MM-GBSA predictive binding energy calculations, and molecular dynamic simulation15,754 natural and synthetic compoundsMain proteaseCompound 2 (molecular bank code AAA396) and compound 3 (molecular bank code AAD146)Chunduru et al. [228]2021In silicoMolecular dockingNovel drug-like inhibitors for COVID-19Main proteaseStructure 61 was found to be more stable and can be further assessed for their antiviral activity to combat COVID-19Coelho et al. [229]2020In silicoBiochemical high-throughput screeningCompound library containing known drugs, bioactive molecules, and natural productsMain proteaseOrganomercuric compounds thimerosal and phenylmercuric acetate, benzophenone, Evans blue, a sulfonic acid-containing dyeDai et al. [230]2020In silicoStructure-based design—Main proteaseDesigned and synthesized two lead compounds (11a and 11b) targeting main protease. Both exhibited excellent inhibitory activityde lima Menezes and da Silva [231]2020In silicoMolecular dynamic simulations, molecular dockingDrugBank databaseNonstructural protein 1 (nsp1)Tirilazad, phthalocyanine, and Zk-806450 showed better energy score than control molecules that have in vitro activity against nsp1 from SARS-CoV-2Tirilazad, phthalocyanine, and Zk-806450Debnath et al. [232]2020In silicoPharmacophores studies, structure-based virtual screening, molecular dynamic (MD) simulationDrug molecule information retrieved from DrugBankMain proteaseDB07456 and DB13592 displayed a similar type of binding interaction with co-ligands and remdesivir, and the predicted Ki values of 2 inhibitors were found to be superior to remdesivirDi Micco et al. [233]2021In silicoMolecular docking, MM-GBSA-based predictions, and molecular dynamicsZonulin inhibitor larazotide acetate (also called AT1001)Main proteaseAT1001, besides its well-demonstrated effect in ameliorating mucosal permeability in ALI/ARDSEl Hassab et al. [234]2021In silicoStructure-based virtual screening, molecular dynamic simulation, and MM-PBSA approaches48 million drug-like compounds of the ZINC databaseSARS-CoV-2 2′-O-methyltransferase (nsp16)Compound 11 as the best potential nsp16 inhibitor herein identified, as it displayed a better stability and average binding free energy for the ligand-enzyme complex compared to sinefunginElmessaoudi-Idrissi et al. [235]2020In silicoIn silico screening, molecular docking, and dynamic approaches5000 compounds of the ZINC databaseMain proteaseThe prominent drug-like and potent inhibitory compounds are 2-[2-(2-aminoacetyl) aminoacetyl] amino-3-(4-hydroxyphenyl)-propanamide (ZINC000004762511), 6′-fluoroaristeromycin (ZINC000001483267), and cyclo(L-histidyl-L-histidyl) (ZINC000005116916) scaffoldsFeitosa et al. [236]2020In silicoMolecular docking—Main proteaseMelatonin can have response potential in early stages for its possible effects on ACE2 and main protease, although it is also promising in more severe stages of the disease for its action against hyper-inflammationDo not confirm antiviral activity, but can rather be used as a basis for further preclinical and clinical trialsGurung et al. [237]2021In silicoVirtual screening, molecular docking1,36,191 moleculesSpike (S) protein receptor-binding domain (RBD) to the host cell surface receptor, angiotensin-converting enzyme 2 (ACE2)ZINC33039472 exhibited binding free energy value lower as compared to the control (emodin) with a higher contribution by gas-phase energy and van der Waals energy to the total binding free energyHaider et al. [238]2020In silicoComputer-aided drug design (CADD) molecular dockingZINCPharmerMain protease∼200 compounds were identified as having strong interaction with Mpro (ZINC20291569, ZINC90403206, ZINC95480156) that showed the highest binding energyHall and Ji [239]2020In silicoHomology modeling molecular dockingZINC15 database: 3447 entriesSpike glycoprotein and main proteaseZanamivir, indinavir, saquinavir, and remdesivir are among the exciting hits of main proteinaseFlavin adenine dinucleotide (FAD) adeflavin, B2 deficiency medicine, and coenzyme A, a coenzyme may also be potentially used for the treatment of SARS-CoV-2 infectionsHavranek and Islam [240]2020In silicoDocking and molecular dynamics2692 protease inhibitor compoundsMain proteasePhenyltriazolinones (PubChem ID: 104161460) and allosteric HCV NS5B polymerase thumb pocket 2 (PubChem ID: 163632044) have shown antiviral activity and also have high affinity towards the main proteaseIbrahim et al. [241]2021In silicoMolecular dynamic simulations, molecular docking, MM-GBSA18 anti-COVID-19 drug candidates against SARS-CoV-2 main proteaseMain proteasePromising binding affinities of TMC-310911 and ritonavirJaiswal and Kumar [242]2020In silicoDocking studies and molecular dynamic simulationDesigned a protein inhibitorSpike (S) glycoproteinThe proposed inhibitor ΔABP-D25YJamalan et al. [243]2021In silicoDocking and molecular dynamic (MD) simulationVirtual screening based on GRL-0617Papain-like proteinase5-(aminomethyl)-2-methyl-N-[(1R)-1-naphthalen-1-ylethyl]benzamide outperformed GRL-0617 in terms of binding affinity (−9.7 kcal/mol). 2-(4-fluorobenzyl)-5-nitro-1H-isoindole-1,3(2H)-dione, 3-nitro-N-[(1r)-1-phenylethyl]-5-(trifluoromethyl)benzamide, 5-acetamido-2-methyl-N-[(1S)-1-naphthalen-1-ylethyl]benzamideJin et al. [244]2020In silicoStructure-assisted drug design, virtual drug screening, and high-throughput screening10,000 compoundsMain proteaseEbselen, disulfiram, tideglusib, carmofur, shikonin, PX-12Ebselen also exhibited promising antiviral activity in cell-based assaysKanhed et al. [245]2021In silicoSystematic virtual screening approachASINEX BioDesign library approved drug libraryMain proteaseRitonavir, nelfinavir, and saquinavir were predicted to be the most potent Mpro inhibitors. 20 molecules (pyrazoles, cyclic amides, pyrrolidine-based compounds, and miscellaneous derivatives)Kavitha et al. [246]2020In silicoMolecular docking molecular dynamic simulations1000 protease-inhibitor-like compounds available in the ZINC databaseMain protease1,2,4 triazolo[1,5-a] pyrimidin-7-onesKrishnan et al. [247]2020In silicoMolecular docking3978 compounds with potential antiviral activityEndoribonuclease (NSP15)8 compounds with good docking score and docking energy e.g., Z595015370, Z1343129850, and Z2760938911Kumar et al. [248]2021In silicoMolecular docking molecular dynamic simulation molecular mechanic Poisson–Boltzmann surface area approachesMillion molecules and natural compound databasesMain proteaseThree compounds namely ZINC14732869, ZINC19774413, and ZINC19774479 displayed better binding affinitiesKumar et al. [249]2020In silicoMolecular docking molecular dynamic simulations13 approved antiviral drugsMain proteaseIndinavir was described as a lead drug. Indinavir possesses an important pharmacophoreNovel compound 16(hydroxyethylamine derivative) suitability as a strong candidate for therapeutic discovery against COVID-19Kwarteng et al. [250]2020In silicoBioinformatic approach molecular docking molecular dynamic simulations—Nucleocapsid (N) proteinZidovudine triphosphate, an anti-HIV agent, as a potential inhibitor of the N-terminal domain of SARS-CoV2 N proteinLi et al. [251]2021In silicoMolecular docking21 antiviral, antifungal, and anticancer compoundsPapain-like proteaseNeobavaisoflavoneMathpal et al. [252]2020In silicoMolecular docking molecular dynamic simulation MM-PBSA3180 FDA-approved drugs from “the ZINC database”Main proteaseZINC03831201, ZINC08101052, ZINC01482077, and ZINC03830817Maurya et al. [253]2020In silicoMolecular dockingAntiviral, anti-infectious, and anti-protease compoundsNSP10/NSP16 methyltransferase and main proteaseCyclocytidine hydrochloride, trifluridine adonitol, and meropenem penciclovir bound with a good docking score NSP10/NSP16 methyltransferase complexed with telbivudine, oxytetracycline dihydrate, methyl gallate, 2-deoxyglucose, and daphnetinMohamed et al. [254]2020In silicoMolecular docking12 histone deacetylases (HDACs)Main proteaseRomidepsin and its active form (RedFK)Monajemi and Zain [255]2021In silicoONIOM (own N-layered integrated molecular orbital and molecular mechanics; QM/MM) approachN3, ebselen, disulfiram, tideglusib, carmofur, shikonin, and PX-12Main proteaseN3, ebselen, and PX-12 inhibitorsBetter inhibition from PX-12 than ebselenMotiwale et al. [256]2020In silicoMolecular docking molecular dynamic simulationsPreviously reported SARS-3CL protease inhibitorsMain proteaseN-substituted isatin derivatives and pyrazoloneMutlu et al. [257]2020In silicoStructure-based approach molecular docking molecular dynamic simulationsFDA-approved and investigational drugsNsp12/Nsp8Two drugs, RX-3117 (fluorocyclopentenyl cytosine) and nebivololNaidoo et al. [258]2020In silicoMolecular docking molecular dynamic simulation MM-PBSACyanobacterial metabolitesMain protease (Mpro) and the papain-like protease (PLpro)Deoxycylin, drospermopsinCylindrospermopsin, deoxycylindrospermopsin, carrageenan, cryptophycin 52, eucapsitrione, tjipanazole, tolyporphin, and apratoxin A exhibited promising inhibitory potential against the SARS-CoV-2 Mpro. The compounds cryptophycin 1, cryptophycin 52, and deoxycylindrospermopsin were observed to display encouraging binding energy scores with the PLpro of SARS-CoV-2Olubiy et al. [259]2020In silicoMolecular docking molecular dynamic simulationsApproved drugs, investigational drugs, natural products, and organic compoundsMain proteaseSeveral tyrosine kinase inhibitors, which include a bioflavonoid and steroid hormones, bind best to main proteaseNilotinib, enasidenib, afatinib, ertapenem, phthalocyanine, hypericin, amrubicin, theacitrin A, theaflavin, amentoflavone, epigallocatechin gallate, glabrolide, cortisol, estradiol, testosteroneÖzdemir et al. [260]2020In silicoMolecular docking density functional theory (DFT) ADME-Tox42 coumarin derivativesMain protease6,7-Dihydroxy-3-phenylcoumarin derivatives gave relatively higher scores, and for all coumarins, and 4-trifluoromethylphenyl substituted coumarin had the highest scoreOzdemir et al. [261]2020In silicoMolecular docking molecular mechanics Poisson–Boltzmann surface area (MM-PBSA)Coumarin derivativesSpike S1 subunit, NSP5, NSP12, NSP15, and NSP16The highest score (−10.01 kcal/mol) in the coumarin group is 2-morpholinoethan-1-amine substituted coumarinPatel et al. [262]2021In silicoPharmacophore studiesDrug library (having drugs and diagnostic agents, which are approved by FDA or other world authorities) and the ASINEX BioDesign libraryMain proteaseRitonavir, nelfinavir, and saquinavir were predicted to be the most potent Mpro inhibitors 20 molecules categorized into four classes viz. disubstituted pyrazoles, cyclic amides, pyrrolidine-based compounds, and miscellaneous derivativesPeng et al. [263]2020In silicoDrug repositioning through virus-drug association prediction—Against SARS-CoV-2Ribavirin was predicted to be the best small molecular drug, with a higher molecular binding energy with human angiotensin-converting enzyme 2 (ACE2), followed by remdesivir, mycophenolic acid, and chloroquine (−6.29 kcal/mol)Pratama et al. [264]2020In silicoMolecular dockingNovel 5-O-benzoylpinostrobin derivativesMain proteaseThree 5-O-benzoylpinostrobin derivatives each with fluoro, tertiary butyl, and trifluoromethyl substituents at 4-position of benzoyl group showed the lowest free energy of binding value and the highest similarity of ligand-receptor interactions with co-crystallized ligandsPundir et al. [265]2020In silicoPharmacophore-based virtual screening molecular mechanics Poisson–Boltzmann surface area (MM-PBSA)PubChem databaseMain proteaseTwo compounds: PubChem3408741 and PubChem4167619 had the binding free energy of −94.02 kJ·mol−1 and −122.75 kJ·mol−1, respectively, as compared to reference X77 (−76.48 kJ·mol−1)Lead molecules for targeting Mpro enzymeQuimque et al. [266]2020In silicoMolecular docking molecular dynamic simulation ADME properties97 antiviral secondary metabolites from fungiPapain-like protease, RNA-dependent RNA polymerase, main protease, spike glycoprotein, nonstructural protein 15 (nsp15)Two fumiquinazoline alkaloids quinadoline B (19), scedapin C (15), and polyketide isochaetochromin D1 (8)Quinadoline B [19] was predicted to confer favorable ADMET values, high gastrointestinal absorptive probability, and poor blood-brain barrier crossing capacitiesRakib et al. [267]2021In silicoMolecular docking molecular dynamic simulation ADME propertiesSelenium-containing heterocyclic compoundsMain proteaseSelection of the 16 most effective selenocompounds as potential anti-COVID-19 drug candidates. Ethaselen showed potential binding affinitiesRane et al. [268]2020In silicoMolecular docking molecular dynamic simulationsDiarylpyrimidine analogsSpike glycoproteinAP-NP (2-(2-amino-5-(naphthalen-2-yl)pyrimidin-4-yl)phenol), AP-3-OMe-Ph (2-(2-amino-5-(3-methoxyphenyl)pyrimidin-4-yl)phenol), and AP-4-Me-Ph (2-(2-amino-5-(p-tolyl) pyrimidin-4-yl)phenol) from a group of diarylpyrimidine derivatives, which appears to bind at the interface of the hACE2-S complex with low binding free energyRao et al. [269]2020In silicoMolecular docking molecular dynamic simulationsVarious small moleculesMain proteasePyranonigrin A, a secondary fungal metaboliteSalman et al. [270]2020In silicoMolecular docking molecular dynamic simulation ADME propertiesLibrary of immunomodulatory medicinal compounds with antiviral capabilitySARS proteases, spike protein, and nonstructural proteins (NSP-9, 15)6 compounds: arzanol, ferulic acid, genistein, resveratrol, rosmanol, and thymohydroquinoneGood pharmacokinetic properties and low acute toxicity of these compoundsSarma et al. [271]2020In silicoMolecular docking MM-GBSA binding free energy molecular dynamic simulations56,079 compounds from ASINEX and Maybridge libraryRNA-binding N-terminal domain (NTD) of the N proteinZINC00003118440 is a theophylline derivative. Pyrimidone derivatives as possible inhibitors of RNA binding to the N-terminal domain of N protein of coronavirusLead moleculesSepay et al. [272]2020In silicoMolecular docking, bioinformatics, and molecular electrostatic potential ADME studiesBenzylidenechromanones, naturally occurring oxygen heterocyclic compoundsMain protease(Z)-3-(4 ′-chlorobenzylidene)-thiochroman-4-Effective pharmacological propertiesShehroz et al. [273]2020In silicoPharmacophore modeling molecular dockingDrugBank, ZINC, and TIMBLE databasesSpike (S) proteinOnly eight molecules fit the criteria for safe oral drugsLead moleculesSingh and Das [274]2021In silicoMolecular dockingChloroquine (CQ) hydroxychloroquine (HCQ) azithromycinSpike (S) protein main protease host cathepsin L (CTSL) receptor-binding domain (RBD)Azithromycin affinity scores (ΔG) with strong interactions with ACE2, CTSL, Mpro, and RBD. CQ firm bond score with Mpro HCQ and two results (ACE2 and Mpro) were firmly bound to the receptorsStefaniu et al. [275]2020In silicoMolecular docking density functional theory (DFT) computations, drug-likeness assessmentDerivatives of benzoic acidMain protease2,5-dihydroxybenzoic acid (gentisic acid) and octylA combination of the two compounds can be consideredTachoua et al. [276]2020In silicoMolecular docking and structural dynamic study molecular mechanic Poisson–Boltzmann surface area (MM-PBSA) ADMET analysisChloroquine, quinine, nitazoxanide, doxycycline, lymecycline, cetirizine, mizolastine, indinavirMain proteaseLymecycline mizolastineUniyal et al. [277]2020In silicoStructure-based virtual screening molecular docking MM-GBSA binding free energy molecular dynamic simulationsCommercially available chemical librariesMain proteaseCompound AG-690/11203374_1 and AG-690/11203374_2 emerged as the best in silico hits based on the docking, MM-GBSA, dynamics, and ADMET studiesLead moleculesWelker et al. [278]2020In silico/in vitroStructure-activity relationships molecular docking fluorescence-based enzyme‐activity assay Vero E6 cellsA series of rationally designed competitive, noncovalent, nonpeptidic active site‐directed SARS‐CoV PLpro inhibitorsPapain-like cysteine proteases (PLpro)R)‐5‐amino‐2‐methyl‐N‐(1‐(naphthalen‐1‐yl)ethyl) benzamide (2b), which is known to bind into the S3 and S4 pockets of the SARS‐CoV PLpro. Isoindoline as a new class of potent PLpro inhibitorsIC50 value of 2.9 ± 0.2 μMWen et al. [279]2021In silicoStructure-based screening8,820 compoundsMain proteaseTrichostatin AA histone deacetylase inhibitor and an antifungal compoundWhite et al. [280]2020In silicoHomology modeling and molecular dynamics approach molecular docking∼970,000 chemical compoundsHelicase (Nsp13)Nilotinib and lumacaftor have significant activity in inhibiting purified recombinant SARS-CoV-2 helicaseWu et al. [281]2020In silicoMolecular docking11 HIV-1 protease inhibitors, 12 nucleotide-analog inhibitors, 728 approved drugsMain protease RNA-dependent RNA polymeraseRemdesivir shows the best binding energy on RdRp and saquinavir is the best inhibitor of main proteaseZaher et al. [282]2020In silicoDesign, synthesis SAR study molecular dockingNewly synthesized sixteen halogenated triazole compoundsHelicase (Nsp13)The most potent compounds were 4-(cyclopent-1-en-3-ylamino)-5-(2-(4-iodophenyl)hydrazinyl)-4H-1,2,4-triazole-3-thiol [16] and 4-(cyclopent-1-en-3-ylamino)-5-[2-(4-chlorophenyl)hydrazinyl]-4H-1,2,4-triazole-3-thiol [12]Zarezade et al. [283]2021In silico3D-QSAR pharmacophore modeling ADMET properties, molecular docking molecular dynamic simulation MM-PBSA, hybrid QM-MM method de novo ligand designPubChem and ZINC databasesHuman angiotensin-converting enzyme 2 and main proteaseZINC12562757 and 112,260,215 were the best potential inhibitors of the ACE2 and main protease, respectively. Evo_1 compound enjoys the highest docking energy among the designed de novo ligandsEvo_1 has a stronger potential for specific inhibition of main protease, as compared to the 112, 260, 215 compound. Lead moleculesIn silico studies used the following component of novel coronavirus as targets: main protease (N = 154), spike glycoprotein (N = 62), nonstructural protein (N = 45), RNA-dependent RNA polymerase (N = 21), papain-like protease (N = 19).About 260 drugs were repurposed by the computational methods for COVID-19 therapy such as about 120 drugs candidate against the main protease, 52 drugs against the spike glycoprotein, 14 drugs against RNA-dependent RNA polymerase, and 28 drugs against other nonstructural proteins.Among the studied repurposed drugs, the best results (regarding efficacy) were observed with saquinavir (N = 9 study), ritonavir (N = 8 study), lopinavir (N = 6 study), remdesivir (N = 3 study), and amikacin, danoprevir, favipiravir, and telaprevir.Table1 shows target-based synthesis of data for COVID-19 drug repurposing. As presented, at least two studies show the efficacy of aprepitant, cobicistat, dipyridamole, and dihydroergotamine against the main protease and tegobuvir (N = 2) against spike protein. The following list of drugs had multitarget action: avapritinib, famotidine, bictegravir, ziprasidone, capmatinib, pexidartinib, amprenavir, zafirlukast, cilostazol, paromomycin, lopinavir, and remdesivir.A total of 91 studies used in silico methods to evaluate the effects of natural products including herbal medicine against SARS-CoV-2. Among them, 54 studies used the main protease as main target, in which eucalyptus (N = 3), Tinospora cordifolia (N = 3), and flavonoids (e.g., hesperidin, rutin, and herbacetin) were the most studied and effective. Other studied targets were as follows: spike (N = 22) and multitarget (N = 20). From the plant metabolites, oleanolic acid, hesperidin, epigallocatechin gallate, jensenone, tinosponone, and anistone show promising results in computational methods against COVID-19. Aloe, green tea, eucalyptus, curcumin, and many Chinese, Indian, and African plants were also effective on COVID-19 in silico.The derivatives of pyrazoles, oxadiazoles, phenyltriazolinones, triazoles, benzoylpinostrobin, benzoic acid, benzylidenechromanones, coumarin, and selenium show efficacy in computational methods and could be considered as lead molecules for drug design and synthesis against COVID-19. Melatonin, ebselen, phenyl furoxan, thimerosal, isatin, romidepsin, phenyl mercurin, pleconaril, and tyrosine kinase inhibitors (such as nilotinib and imatinib) are examples of other drugs that are effective in in silico methods.


## 3.2. Results from In Vitro and In Vivo Studies

Screening of in vitro studies leads to finding 34 studies. Different compounds were evaluated and most studies focused on the inhibition of viral replication, which was assessed by the quantification of viral RNA by PCR and IC50 value reported. Most of the studies use the Vero E6 cell line for the assessment of replication of SARS-CoV-2 (Table 4). In particular, main protease, papain-like protease, RNA-dependent RNA polymerase, NSP-14, NSP-15, spike protein, and TMPRSS2 were evaluated in vitro as targets. Also, three studies focused on in vitro inhibition of inflammatory markers such as interleukins and the effects of the cytopathic effect on infected cell.Table 4
Summary of studies with in vitro method.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsAgarwal et al. [284]2020In vitroIL-1β assay using THP-1 cells, in vivo pharmacokinetic investigation in C57BL/6 miceAlkenyl sulfonylurea derivativesNLRP3 inhibition and reduction in the release of interleukin-1βIL-1β inhibition IC50 of 35 nM, good oral absorption showing Cmax of 8.49 μg/mL with an AUC of 48.9 μg·h/mL and terminal half-life of 2.86 h, after oral route of administration at 3 mg/kg doseNovel thiazolo-alkenyl sulfonylurea derivative7 as potent, selective, and orally bioavailable NLRP3 inflammasome inhibitorAkaberi et al. [285]2020In vitroVero E6 cells, RT-qPCR for the quantification of viral RNA, expression and purification of SARS-CoV-2 3CL protease in vitro enzymatic assayNO-Donor S-nitroso-N-acetylpenicillamine (SNAP): nitric oxideMain proteaseAlthough the viral replication was not completely abolished (at 200μM and 400 μM), SNAP delayed or completely prevented the development of viral cytopathic effect in treated cells (IC50 = 440.95 μM ± 36.15 SE)A dose-dependent inhibition of the 3CL protease (400μM)Bernstein and Zhang [286]2020In vitroVero E6 cells: virus was then added at a multiplicity of infection (MOI) of 0.01, with 1 h allowed for infection. Cytotoxicity: Cell Counting Kit-8 (CCK-8) colorimetric assayGallium maltolate (GaM)Inhibition of viral replicationEC50 (concentration producing 50% inhibition of viral replication) of about 14 μM (CI 95%: 8.9–22.8 μM)No cytotoxicity was observed at concentrations up to at least 200μMBocci et al. [287]2020In vitroLigand-based virtual screening/Vero E6 cells. Uninfected cells and chloroquine as control inhibition of the SARS-CoV-2-mediated CPE following infection in Vero E6 cells tissue culture infectious dose (TCID50) assay4000 approved drugs/glafenine, amodiaquine vorinostat, zuclopenthixol, isoxsuprine, nebivolol, ambroxol, panobinostat, and pracinostatInhibition of viral replicationIn silico: glafenine, amodiaquine (amodiaquine), vorinostat, zuclopenthixol, isoxsuprine, nebivolol, ambroxol, panobinostat, and pracinostat. In vitro: zuclopenthixol and amodiaquine show anti-SARS-CoV-2 activity comparable with chloroquine. EC50 values were estimated as 0.13 μM for AQ, 1.35 μM for ZPX, and 2.72 μM for nebivololZuclopenthixol and nebivolol exhibitin vitro antiviral activities and potencies that are comparable or better than chloroquine and hydroxychloroquineCao et al. [288]2020In vitroAfrican green monkey kidney Vero E6 cells were infected with virus at an MOI of 0.01 for 1 h. Immunofluorescence assay physiologically based pharmacokinetic modeling and simulations9 artemisinin-related compoundsInhibition of viral replicationArteannuin B showed the highest anti-SARS-CoV-2 potential with an EC (50) of 10.28 ± 1.12μM. Artesunate and dihydroartemisinin showed similar EC (50) values of 12.98 ± 5.30 μM and 13.31 ± 1.24 μM, respectivelyLumefantrine could inhibit SARS-CoV-2in vitro with an EC50 of 23.17 ± 3.22 μM. Impair viral infection by modulating host cell metabolic pathwaysChen et al. [289]2021In vitroFluorescence-based high-throughput screen, docking/pretreated with various concentrations of test compound for 1 h, followed by infection with SARS-CoV-2 (MOI of 0.0001) in the presence of test compounds, expression and purification of SARS-CoV-2 PLpro, 3CL protease fromEscherichia coli enzymatic assay of SARS-CoV-2 PLpro cytotoxicity assay1920 natural productsMain protease, papain-like proteaseGinkgolic acid and anacardic acid were also identified as inhibitors of 3CLpro, with IC50 values of 1.79 ± 0.58 and 2.07 ± 0.35 μM, respectively, both ginkgolic acid and anacardic acid dose-dependently inhibited PLpro activity, with IC50 values of 16.30 ± 0.64 and 17.08 ± 1.30 μM, respectivelyGinkgolic acid act as an irreversible inhibitor against both PLpro and 3CLpro, suggesting it is a covalent inhibitorGendrot et al. [290]2020In vitroVero E6 cells, replication was estimated by RT-PCR/chloroquine as control determination of the inhibition stageDoxycyclineInhibition of viral replicationMedian effective concentration (EC50) and 90% effective concentration (EC90) for doxycycline were 4.5 ± 2.9 μM and 23.5 ± 16.5 μM, respectively, EC50 = 4.5 μMInteracted at both entry and postentry stages of the SARS-CoV-2 infection doxycycline has anti-inflammatory effectsGupta et al. [291]2021In vitroHigh-throughput virtual screening, molecular dynamic simulation, prime MM-GBSA/Vero E6 cells infected with the SARS-CoV-2 fluorescence-based biochemical assay for inhibitors of 3CLproFDA-approved drugsMain protease, PLpro, RNA-directed RNA polymerase, helicase, NSP-15, NSP14, and 2′-O-methyltransferaseTroxerutin (main protease and PLpro) and bisindolylmaleimide derivatives (main protease and exon) had possible dual targets. Ivermectin has nonselective toxicity to the ATCC E6 Vero cells at ≤50μM and 16.67 μM based on the number of nuclei countedBIM IX specifically blocked 3CLproin vitro enzymatic assay inhibition was observed with an IC50 value of 113.7 ± 5.2 μM a known inhibitor of protein kinase C isoforms, bisindolylmaleimide IX (BIM IX), was found to be a potent inhibitor of SARS-CoV-2Hahn et al. [292]2020In vitroVero B4 and 76 cells و Vero E6 cells/remdesivir RT-qPCR for the detection of extracellular SARS-CoV-2 in cell immunostaining for the detection of Intracellular SARS-CoV-2 Western blot analysis viral plaque and yield reduction assay neutral red assay (NRA)(IMU-838) inhibitor of human dihydroorotate dehydrogenase (DHODH)Inhibition of viral replication/immunomodulatorA virus-specific EC90 of 6.2 ± 1.9 μM was measured, with no cytotoxicity observed with drug concentrations up to 100 μM at 3 days p.i.Active moiety of IMU-838, vidofludimus, possesses broad-spectrum antiviral activity. IMU-838 reduces T lymphocyte proliferation, cytokine production, and organ infiltration by leukocytes in various in vivo and in vitro models for autoimmunityHattori et al. [293]2021In vitroVeroE6 cell-based assays with RNA-qPCR, cytopathic assaysTwo small-molecule-compounds, named GRL-1720 and 5 h, containing an indoline and indole moietyMain proteaseEC(50) values of 15 ± 4 and 4.2 ± 0.7μM for GRL-1720 and 5 hCombination of 5 h and remdesivir exhibits synergism against SARS-CoV-2.5 h might serve as a lead M (pro) inhibitorJang et al. [294]2021In vitroVero CCL-81 cells/Calu-3 human lung epithelial cellsGemcitabine and its analog 2′-fluoro-2′-deoxycytidine (2FdC)Inhibition of viral replication50% effective concentration (EC(50)) of 1.2μM compare to remdesivir 2FdC was marginally active (EC(50) = 175.2 μM)Gemcitabine has a synergistic effect when combined with remdesivirJeon et al. [295]2020In vitroDrug repositioning/chloroquine, lopinavir, and remdesivir were used as reference drugs/VeroE6 cell/immunofluorescence analysis48 FDA-approved drugsInhibition of viral replicationNiclosamide (IC50, 0.28 μM), ciclesonide (IC50, 4.33 μM)24 potential antiviral drug candidates against SARS-CoV-2 infectionLiu et al. [296]2020In vitroVirtual screening/spiked pseudotyped lentivirus for imitating SARS-CoV-2 cell entry/ACE2-GFP expressing HEK293T 2 cells. HeLa Cells, 293T (human, kidney) cells, and Vero-E6 (African green monkey, kidney) cellsFDA-approved drugsAngiotensin I-converting enzyme 2 (ACE2) and a spike proteinNine potential candidates were selected and submitted to experimental studies. Three (romidepsin, saquinavir, and nelfinavir) of nine drugs possess the ability to suppress SARS-2-S pseudotyped particles to enter the ACE2 expressing cells in a concentration-dependent mannerFive clinical HDAC inhibitors including romidepsin, panobinostat, givinostat hydrochloride monohydrate, CAY10603, and sirtinol could inhibit noticeably the spike/ACE2-mediated cell entry of SARS-CoV-2Ma et al. [297]2020In vitroVero E6 cells using cytopathic effect and plaque reduction assay. HCoV-229E infection in Huh-7 cells. Western blot assayPhillyrin (KD-1) ingredient of Forsythia suspensaInhibition of viral replicationInhibit SARS-CoV-2. Markedly reduce the production of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β, MCP-1, and IP-10) at the mRNA levels. p-NF-κB p65, NF-κB p65, and p-IκBαKD-1 could significantly inhibit virus proliferation in vitro, the upregulated expression of pro-inflammatory cytokines induced by SARS-CoV-2Masih et al. [298]2020In vitroLPS-stimulated RAW267.4 cells by enzyme immunoassay Western blot assayPyrazole derivativesAnti-inflammatory levels of interleukin-1β, tumor necrosis factor-α, and interleukin-6Compound 6c as the most potent analog among the tested series. Inhibition of inhibitor kappa B-α and NF-κBOmotuyi et al. [299]2020In vitroKinetic assayAframomum melegueta. 100 secondary metabolitesMain protease, 2′-O-ribose methyltransferase (NSP16), and surface glycoprotein/ACE2 receptor interfaceDiarylheptanoid (letestuianin A), phenylpropanoid (4-cinnamoyl-3-hydroxy-spiro[furan-5,2′-(1′H)-indene]-1′,2,3′(2′H,5H)-trione), flavonoids (quercetin, apigenin, and tectochrysin) have been identified as high binding compounds to SARS-CoV-2 targets in a polypharmacology mannerDi-ethyl-ether (IC50 = 0.03 mg/L), acetone (IC50 = 1.564 mg/L), ethyl-acetate (IC50 = 0.382 mg/L), and methanol (IC50 = 0.438 mg/L) fractions demonstrated the best inhibition in kinetic assayOutlaw et al. [300]2020In vitroVero E6 cellβ-Gal complementation-based fusion assay. Viral titration and plaque reduction neutralization assay quantitative RT-PCR.A lipopeptide that is derived from the C-terminal heptad repeat (HRC) domain of SARS-CoV-2 SInhibition cell-cell fusionInhibits cell-cell fusion mediated by SARS-CoV-2 S and blocks infection by live SARS-CoV-2 in Vero E6 cell monolayers (IC50) of ∼10 nM and 90% inhibitory concentration (IC90) of ∼100 nMPitsillou et al. [301]2021In vitroMolecular docking, molecular dynamic simulation/papain-like protease (SARS-CoV-2) assay kit GRL-0617 as a control enzymatic inhibition assay300 small compounds derived predominantly from our OliveNet™ library (222 phenolics) and supplemented with synthetic and dietary compoundsPapain-like proteaseSelection of 30 compounds. Hypericin possessed inhibition activity, and both rutin and cyanidin-3-O-glucoside resulted in a concentration-dependent inhibition of the PLproThe IC50 values were not clarified in experimentPizzorno et al. [302]2020In vitroVero E6 cells/remdesivir, lopinavir, chloroquine as comparatorFavipiravir, ribavirin, umifenovir (arbidol), berberine, cyclosporine A, diltiazemInhibition of viral replicationUmifenovir, berberine, and cyclosporine A with estimated 50% inhibitory concentrations of 0.99, 5.2, 1.38, 3.5, 10.6, and 3μM, respectivelyA strong antagonism between remdesivir and berberine, in contrast with remdesivir/diltiazem, for which we describe high levels of synergyPlaze et al. [303]2021In vitroMonkey VeroE6 cells and human alveolar basal epithelial A549-ACE2 cell RT-qPCR for the presence of SARS-CoV-2 RNAChlorpromazineInhibition of viral replication(IC50) of 8.2 μM in monkey VeroE6 cells (CC50) of 13.5 μM, and selectivity index (SI) of 1.65 IC50 of 11.3 μM, CC50 of 23.1 μM, and SI of 2.04 in human A549-ACE2 cellsReznikov et al. [304]2021In vitroRepurposing study molecular docking/human lung A549 cells that were transfected with hACE2 (ACE2-A549) cells, Vero E6 cellsElectronic health records of over 219,000 patients with antihistaminesInhibition of viral replication/spike proteinIC50 15.3 μg/ml for hydroxyzine, 17.4 μg/ml for diphenhydramine, and 2.24 μg/ml for azelastineHydroxyzine and possibly azelastine bind angiotensin-converting enzyme-2 (ACE2) and the sigma-1 receptor as off-target disease preventionSakurai et al. [305]2021In vitroVeroE6 cells/human colon-derived Caco-2 cells/infection assay with immunofluorescence5-Amino levulinic acid (5-ALA)Inhibition of viral replicationIC50 of 570 μM/IC50 of 63 μM in human Caco-2 cells. No significant cytotoxicity5-ALA is synthesized in most animals and plants and we are continuously consuming it in our food. Show anti-inflammation effects in humansStone et al. [306]2021In vitroVero E6 monkey kidney and Calu-3 human lung adenocarcinoma cells/RT-qPCR of RNAStenoparib poly(ADP ribose) polymerase (PARP) inhibitorInhibition of viral replicationDose-dependent activity against SARS-CoV-2 at concentrations up to 30μM with negligible cytotoxicity. Synergistic effect with remdesivirStenoparib impedes entry and postentry processesSu et al. [307]2020In vitroVero E6 cells, quantitative real-time RT-PCRShuanghuanglian preparation, a Chinese traditional patent medicine (chlorogenic acid, phillyrin, baicalin, and baicalein)Main proteaseDose-dependent inhibition of SARS-CoV-2 3CLpro, and the resulting IC50 values were 0.090, 0.064, and 0.076 μL/mL for three Shuanghuanglian oral liquids produced by three different pharmaceutical companiesBaicalin and baicalein, which have good drug-like propertiesTouret et al. [308]2020In vitroVeroE6 cells/human colon-derived Caco-2 cell real-time RT-PCR/remdesivir as controlFDA-approved chemical libraryInhibition of viral replication23 drugs were selected to cover the 12 different groups. 11 compounds such as macrolide antibiotics, proton pump inhibitors, antiarrhythmic agents, or CNS drugs emerged showing antiviral potency with 2 < EC50 ≤ 20μMTwo of the highest antiviral activity were obtained for azithromycin (EC50 = 2.12μM) and hydroxychloroquine (EC50 = 4.17 μM). Vonoprazan and sulfadoxine show a moderate activity with EC50 above 30 μMTree et al. [309]2021In vitroPlaque inhibition assay with Vero E6 cells/differential scanning fluorimetry/ELISA assaysUnfractionated heparin (UFH), low MW heparinsInhibition of viral replication/spike protein RBDAll the UFH preparations had potent antiviral effects, with IC50 values ranging between 25 and 41 μg·ml−1, whereas LMWHs were less inhibitory by ∼150-fold (IC50 range 3.4–7.8 mg·ml−1)Heparin directly inhibits the binding of RBD to the human ACE2 protein receptorVatansever et al. [310]2021In vitroDocking, screening assay/Vero E6, and A549 cell culture30 FDA/EMA-approved drugsMain proteaseSix FDA/EMA-approved drugs can potently inhibit Mpro with an IC50 value lower than 100 μM. Bepridil exhibited strong inhibition of SARS-CoV-2 from entry and replication inside Vero E6 and A549 cells (IC50 = 72 μM)Bepridil indicated that it had low micromolar EC50 valuesWang et al. [311]2020In vitroFRET-based enzyme activity assay of Mpro measurement of human TMPRSS2 activity by a FRET-based enzymatic assay surface plasmon resonance (SPR) analysis molecular docking African green monkey kidney cell Vero E6Catechin, kaempferol, quercetin, proanthocyanidins, resveratrol, and tannic acidTMPRSS2 (transmembrane protease serine 2) main proteaseTannic acid inhibited the activities of the two proteases with an IC50 of 13.4 mM for Mpro and 2.31 mM for TMPRSS2A potent dual inhibitorWatashi [312]2021In vitroVirtual drug screening, docking/VeroE6 cells detecting the viral RNA by real-time RT-PCR or viral proteins by immunofluorescenceFDA-approved drugsMain protease, PLpro, RdRp, helicase, spike, ACE2, and TMPRSS2Abiraterone, amodiaquine, anidulafungin, arbidol, astemizole, atazanavir, auranofin, azithromycin bazedoxifene, bexarotene, camostat IC50 values <5 μMCepharanthine, cetilistat, chloroquine, ciclesonide, cyclosporine A, digoxin, ivermectin, lopinavir, mefloquine, nelfinavir, niclosamideYu et al. [313]2021In vitroComputer-aided drug design and biological verification surface plasmon resonance (SPR) assays and NanoBiT assaysTraditional Chinese medicineSpike proteinsGlycyrrhizic acid, the most efficient and nontoxic broad-spectrum anticoronavirus molecule IC50, was 22 μMZandi et al. [314]2021In vitroAfrican green monkey kidney cells (Vero CCL-81 cells) and reverse transcription-quantitative PCR (qRT-PCR). Remdesivir andβ-D-N4-hydroxycytidine as positive drug controlsNucleoside analogs/anti-HCV agentsInhibition of viral replicationSofosbuvir and favipiravir demonstrated no antiviral effect against COVID-2Zhang et al. [315]2020In vitroHybrid virtual screening, docking, force field-based simulation/qRT-PCR assay, indirect immunofluorescence assay (IFA) and CCK-8 assay, surface plasmon resonance (SPR) assay/Vero cells1906 approved drugs from TargetMol-approved drug libraryRNA-dependent RNA polymerase (RdRp)Pralatrexate and azithromycin (EC50) values of 0.008 and 9.453 μMA new therapeutic agent pralatrexate against COVID-19 by targeting RdRpZhang et al. [316]2020In vitroAfrican green monkey kidney Vero E6 cell line, quantitative real-time PCR (RT-PCR) analysis, immunofluorescence microscopy, Western blot analysisLopinavir/ritonavir, rupintrivir, and AG7404Inhibition of viral replication(IC50) and half cytotoxic concentration (CC50) values of lopinavir were ∼12.01 μmol/L and 80.82 μmol/L; for ritonavir, the IC50 and CC50 values were 19.88 μmol/L and 94.71 μmol/L (SI = 4.76)Rupintrivir inhibited SARS-CoV-2 infection only at high drug concentrations (Vero E6: IC50 = 34.08 μmol/L, AG7404: Vero E6: IC50∼195.8 μmol/L). Lopinavir/ritonavir should be stopped for clinical use due to the huge gap between in vitro IC50 and free plasma concentrationZhu et al. [317]2021In vitroHuman fibroblast lung cells (MRC-5), cytopathic effect (CPE) assay, primary human airway air-liquid interface (ALI) cultures, TEER assay, CCK-8 assaysStimulator of interferon genes (STING): dimeric amidobenzimidazole (diABZI)Inhibition of viral replicationPotent anticoronavirus activity against both the common cold human coronavirus 229E (HCoV-229E) and SARS-CoV-2 in cell culture systems. EC50 of 120 μMNitric oxide, ginkgolic acid, anacardic acid, troxerutin, bisindolylmaleimide derivatives, small molecules (GRL-172, and 5 h), baicalin and baicalein (phytochemicals), and bepridil were effective drugs in vitro against the main protease of COVID-19.The following drugs could inhibit the spike/ACE2-mediated cell entry of COVID-19 in vitro: romidepsin, panobinostat, givinostat, sirtinol, saquinavir, lipopeptides, hydroxyzine, azelastine, heparin, and glycyrrhizic acid.Alkenyl sulfonylurea derivatives [7], an IMU-8381 inhibitor of human dihydroorotate dehydrogenase, pyrazole derivatives, and phillyrin, regulated the expression of inflammatory cytokines (e.g., IL, TNF-α, and NF-κB) induced by SARS-CoV-2 markedly and could be considered as adjuvant treatment of COVID-19 severe disease.Doxycycline, chlorpromazine, azithromycin, heparin, bepridil, tannic acid, and glycyrrhizic acid are well-known drugs that show both in silico and in vitro inhibitory effects against SARS-CoV-2 and should be considered for this purpose.Also, in vitro studies show that lopinavir/ritonavir, sofosbuvir, and favipiravir have no antiviral effects against SARS-CoV-2 (huge gap between in vitro IC50 and free plasma concentration).We included 16 in vivo studies in our final analysis. Most of them use a combination of in vitro and in vivo methods for the evaluation of novel drugs. Table5 shows the detail of these studies.Table 5
Summary of studies with in vivo method.
AuthorYearMethodDetail of methodName of compound/drugTargetEfficacyCommentsLei et al. [318]2020In vivoBALB/c mice to determine the pharmacokinetic profiles of the fusion proteinsA recombinant protein by connecting the extracellular domain of human ACE2 to the Fc region of the human immunoglobulin IgG1Spike proteinThe IC50 values of ACE2-Ig for SARS-CoV and SARS-CoV-2 neutralization were 0.8 and 0.1 μg·ml−1, respectively. Desirable pharmacological properties in micePotential applications in the diagnosis, prophylaxis, and treatment of SARS-CoV-2Rathnayake et al. [319]2020In vivoOld male hDPP4-KI mice infected with MERSMA-CoVDipeptidyl and tripeptidyl series of compoundsMain protease7a, 6c, 7e, 7h, and 6j: EC50 values ranging from 0.15 to 0.9 μM in Vero E6 cells, IC50: 0.17 to 0.82 μM. 40% of mice treated with compound 6h survived, and all mice treated with compound 6j were alive at the end of the study6j resulted in the survival of MERSMA-CoV-infected hDPP4-KI miceSchäfer et al. [320]2020In vivoAn immune-competent mouse model of COVID-19 by remodeling the SARS-CoV-2 S RBD at the mACE2-binding interface. Human Ace2-expressing HT1080 cells and Syrian hamster model of SARS-CoV-2Human mAbs (hu-mAbs)Reduction in viral load and prevent infectionNeutralizing hu-mAbs targeting SARS-CoV-2 promotes reduction in viral load and prevent infection in macaques and hamsters effective dose: 5.3 and 16 mg/kg inhibitory concentration (IC)50/IC90 (half-maximal/90% IC) of 4.4/18 to 26/140 ng/mlSchumann et al. [321]2020In vivoFemale C57Bl/6 mice, LPS stimulationMP1032:5-amino-2,3-dihydro-1,4-phthalazinedione sodium saltImmune-modulatingDirect inhibitory activity on human PARP-1 was detected in the low micromolar range (IC50 = 1.55 μM). MP1032 works as a ROS scavenger, reduced the secretion of TNF-α and IL-6. MP1032 pre-treatment significantly reduced plasma cytokine levels compared with vehicle-treated mice by 50% (TNF-α) and 25% (IL-6). Significant reduction in SARS-CoV-2 replicationMP1032 could further attenuate prolonged virus replication by preventing oxidative stress or by limiting ADP ribosylation of the viral nucleocapsid protein via PARP-1 inhibitionShang et al. [322]2021In vivoAfrican green female hACE2 transgenic mice, SARS-CoV-2 injection RNA extraction and qPCR, RNA in situ hybridization assay, histopathological examinationEndosomal acidification inhibitors, including chloroquine, bafilomycin A1, and NH4CLAntiviral actions against SARS-CoV-2(40μM), bafilomycin A1 (100 nM), and NH4CL (12.5 mM) suppressed the replication of SARS-CoV-2 in all cell types chloroquine (60 mg·kg−1) and bafilomycin A1 (0.1 mg·kg−1) markedly reduced virus yields in lung tissues. Alleviated viral pneumonia in hACE2 transgenic miceEndosomal acidification inhibitors exhibited antiviral actions against SARS-CoV-2Sheahan et al. [323]2020In vivoMale and female 20- to 29-week old SPF C57BL/6J (Stock 000664 Jackson Labs) mice, whole-body plethysmography 50, 150, or 500 mg/kg EIDD-2801 2 hr prior to intranasal infection with 5E + 04 PFU of mouse-adapted SARS-CoVRibonucleoside analogβ-d-N4-hydroxycytidineAntiviral efficacy against the three most recently emerged human CoV: SARS-CoV, MERS-CoV, and SARS-CoV-2Bioavailable NHC prodrug (β-d-N4-hydroxycytidine-5′-isopropyl ester), improved pulmonary function and reduced virus titer and body weight lossContinued development of EIDD-2801 as a potent broad-spectrum antiviralTai et al. [324]2021In vivoSprague Dawley rats. (i) HCQ‐IV: 12 rats received a single dose of 0.590 mg HCQ sulfate per animal via IV injection; (ii) HCQ‐IT: 20 rats received a single dose of 0.590 mg HCQ sulfate per animal via intratracheal (IT) administration; and (iii) liposomal HCQ‐IT: 20 rats received a single dose of 0.284 mg liposomal HCQ sulfate per animal via IT administration 0.25, 1, 4, 24, and 72 hours postdose and for tissue/organ samples were 0.25, 4, 24, and 72 hours postdoseInhalable liposomal hydroxychloroquine (HCQ)Pharmacokinetic studyCompared with unformulated HCQ administered intravenously, liposomal HCQ showed higher (∼30-fold) lung exposure, longer (∼2.5-fold) half-life in lungs, but lower blood exposure with ∼20% of peak plasma concentration (Cmax) and 74% of area under the curve from 0 to 72 hours (AUC 0–72) and lower heart exposure with 23% of Cmax and 58% of AUC 0–24 (normalized for dose)Inhalable liposomal HCQ may provide clinical benefit and serve as a potential treatment of COVID-19Tortorici et al. [325]2020In vivoFrom two individuals recovering from severe COVID-19 disease. Surface plasmon resonance (SPR) and flow cytometry. Cell-cell fusion assay using VeroE6 cells/Syrian hamster modelHuman neutralizing antibodies (S2E12 and S2M11)Against SARS-CoV-2IC50 values were 1.2 to 6.6 ng/ml to protect hamsters against SARS-CoV-2 challenge competitively block angiotensin-converting enzyme 2 (ACE2) attachment and that S2M11 also locks the spike in a closed conformation by recognition of a quaternary epitope spanning two adjacent receptor-binding domainsAntibody cocktails for prophylaxis or therapyWahl et al. [326]2021In vivoCoronavirus replication in human lung-only mice (LoM) and histopathologic analysis LoM were administered EIDD-2801 starting 24 h or 48 h post-SARS-CoV-2 exposure and every 12 h thereafterRibonucleoside analogβ-D-N4-hydroxycytidine (NHC): oral prodrug EIDD-2801 (also known as molnupiravir or MK-4482)Inhibited SARS-CoV-2 replicationEIDD-2801 pre-exposure prophylaxis significantly reduced virus titers in the human lung tissues of LoM by over 100,000-fold in two independent experimentsProphylactic administration EIDD-2801 is highly effectiveWang et al. [327]2021In vivoMale hACE2 mice inoculated intranasally with SARS-CoV-2 stock virus rhesus macaque model of SARS-CoV-2 infectionScreening the FDA-approved peptide drug library with LibDockAgainst SARS-CoV-2Ten polypeptide drugs were selected. Dalbavancin showed the strongest inhibitory ability. (EC50) was ∼12 nM. Significant inhibition of SARS-CoV-2 pseudovirion entry into HEK293/hACE2 cells, with an IC50 of ∼53 nM in both mouse and rhesus macaque models, viral replication, and histopathological injuries caused by SARS-CoV-2 infection are significantly inhibited by dalbavancin administrationDalbavancin directly binds to human angiotensin-converting enzyme 2 (ACE2) with high affinity, thereby blocking its interaction with the SARS-CoV-2 spike proteinWeston et al. [328]2020In vivoMice were inoculated intranasally with SARS-CoV20 FDA-approved drugsAgainst SARS-CoV-27 of these inhibit SARS-CoV-2 at non-cytotoxic concentrations. Many of the compounds have IC50 under 10 μM, chloroquine and chlorpromazine did not inhibit viral replication in mouse lungs based on viral titers recovered at 4 dpi neither drug inhibited viral replication in the lungs, but both protected against clinical diseaseWhite et al. [329]2021In vivoA human ACE2-expressing adenovirus to transduce the naturally resistant wild-type BALB/c mice and sensitize them to SARS-CoV-2 infection, K18-hACE2 mouse modeleEF1A inhibitor plitidepsin (aplidin): targeted the eukaryotic translation machineryInhibited SARS-CoV-2 replicationIC90 of 1.76 nM: Vero E6 IC90 of 0.88 Nm: human cell line significantly reduced genomic RNA content reduction of nearly 2 log units in SARS-CoV-2 viral titers in the lungs of the 0.3 mg/kg plitidepsin group, reduction of 2 log units in viral lung titers at day 3, similar to two daily 50 mg/kg doses of remdesivirPlitidepsin as a host-targeted anti-SARS-CoV-2 agent with in vivo efficacyWu et al. [330]2020In vivo10 male C57BL/6 mice related to 200 mg/kg/day GB-2 everyday by oral administration. immunohistochemistry (IHC) assessmentGB-2, the formula from Tian Shang Sheng Mu of Chiayi Puzi Peitian Temple: in Taiwan: the index compound ofCamellia sinensis var. assamica extractThe protein and mRNA expression of ACE2 and TMPRSS2No mouse death and no significant alteration in mouse activity or body weight (IHC) data revealed that the expression levels of both ACE2 and TMPRSS2 were markedly diminished in the GB-2 group compared with the control group 50μg/mL of theaflavin could inhibit protein expression of ACE2 and TMPRSS2GB-2 and theaflavin could act as potential compounds for ACE2 and TMPRSS2 inhibitorsYuen et al. [331]2021Ex vivoHuman lung tissues were treated with the indicated drugs overnight, followed by SARS‐CoV‐2 infection at 2 × 106 PFU/wellSmall molecules targeting the ULK1/Atg1 complex involved in the induction stage of autophagy (ULK1 inhibitor SBI0206965), the ATG14/Beclin1/VPS34 complex involved in the nucleation step of autophagy (class III PI3‐kinase inhibitor VPS34‐IN1), and a widely used autophagy inhibitor that persistently inhibits class I and temporary inhibits class III PI3‐kinase (3‐MA) and a clinically approved autophagy inhibitor that suppresses autophagy by inhibiting lysosomal acidification and prevents the formation of autophagolysosome (HCQ)Target the key cellular factors involved in key steps of the autophagy pathwayNot all the tested autophagy inhibitors suppressed SARS-CoV-2 infection inhibition of class III PI3‐kinase Vps34 downstream of ULK1, in contrast to inhibition of ULK1, which caused a significant decrease in SARS‐CoV‐2 replication (EC50) of HCQ was 19μM, that of Vps34‐IN1 was 0.82 μM Vps34‐IN1 potently inhibited SARS‐CoV‐2 viral replication in normal ex vivo human lung tissue culture in a dose‐dependent mannerClass III PI3-kinase may be a possible target for COVID-19 therapeutic developmentZhang et al. [332]2021In vitro/in vivoPreparation of MAbs from female BALB/c mice and wild-type Balb/c mice were intranasally inoculated with hACE2-encoded adenovirus 5 (Ad5-hACE2) to allow expression of the hACE2 receptor in the lung, followed by intranasal infection with live SARS-CoV-2 3 days laterMouse anti-SARS-CoV-2-neutralizing MAbs: 2H2 and 3C1Spike (S) protein(IC50) of 12 ng/mL effectively treat SARS-CoV-2-infected mice even when administered as late as 24 h post-infectionHuman monoclonal antibodies were the most evaluated drugs (N = 4), promote the reduction in viral load (in vitro), and prevent infection in animal models of SARS-CoV-2.A prodrug of hydroxycytidine (molnupiravir) improved pulmonary function and reduced viral titer in vivo and was introduced as a potential broad-spectrum antiviral agent against SARS-CoV-2.Chloroquine and chlorpromazine did not inhibit viral replication in mouse lungs, but protected them against clinical disease.Dalbavancin shows significant inhibitory ability in both in vitro and in vivo models of COVID-19. The drug binds directly to ACE2 and blocks its interaction with the SARS-CoV-2 spike protein.


## 4. Discussion

Drug development is a multistep process, typically requiring more than five years to assure the safety and efficacy of the new compound. There are several strategies in antiviral drugs for coronaviruses including empirical testing of known antiviral drugs, large-scale phenotypic screening of compound libraries, and target-based drug discovery. To date, an increasing number of drugs have been shown to have anticoronavirus activities in vitro and in vivo, but only remdesivir and several neutralizing antibodies have been approved by the US FDA for treating COVID-19. However, remdesivir’s clinical effects are controversial and new antiviral drugs are still urgently needed. Given the urgency of the SARS-CoV-2 outbreak, here we discuss the discovery and development of new therapeutics for SARS-CoV-2 infection, which have been conducted in basic research (in silico) and preclinical study (in vitro, in vivo). Our database search identified about 3000 studies, which means a global effort for drug development in the current COVID-19 pandemic. Although the chance of successful drug development is very low (less than about 10%) and till today, there are no approved drugs. Many potential candidates (at least 420 drugs) are in clinical trials.As summarized in the results, many compounds are in the development process for COVID-19 disease. Some of these compounds are completely new and could serve as seeds (or leads) for developing antiviral drugs against COVID-19, but as we need therapeutics as soon as possible, half of the studies focused on drug repurposing (repositioning), which is a process of investigation of existing drugs for new therapeutic purposes. With the emergence of a growing COVID-19 pandemic, the drug repurposing process was being accelerated. Clinical trials using repurposed drugs may take less time and have a lower overall cost of manufacturing and could have a wide distribution of drugs. According to our results, 260 drugs repurposed by the computational methods for COVID-19, among them saquinavir, ritonavir, and lopinavir, showed the best efficacy in in silico environment. These drugs can be rapidly repurposed for clinic application for treating COVID-19 patients given their proven safety.Many trials are performed using a combination of ritonavir-lopinavir. The results of a systematic review and meta-analysis showed that this drug combination has no more treatment effects than other therapeutic agents in COVID-19 patients and is currently not used anymore [134]. We could not find any clinical trial on saquinavir, which is the most studied drug in silico and show high potency against COVID-19. Saquinavir could be a suggestion for further clinical research.Given that the development of synthetic chemicals for therapeutic use is a random process that might result in serendipitous discovery, many pharmaceutical companies are now focused on the development of plant-derived drugs. Natural products and their structural analogs have historically made a major contribution to pharmacotherapy, especially for cancer and infectious diseases. Nevertheless, natural products also present challenges for drug discovery, such as technical barriers to screening, isolation, characterization, and optimization. In recent years, several technological and scientific developments—including improved analytical tools, genome mining and engineering strategies, and microbial culturing advances—are addressing such challenges and opening up new opportunities. Consequently, interest in natural products as drug leads is being revitalized. Medicinal plants have attracted significant attention to treat infectious diseases. Complex molecular structures and a wide variety of natural compounds make medicinal plants an excellent biological resource for drug discovery. Our results show that various plants have potential antiviral activities and could use or be a basis for drug development against COVID-19.Some of the studied plants are among what used by people on a daily basis, such as green tea, aloe, curcumin, and eucalyptus. A systematic review and meta-analysis of RCTs on herbal medicine in the management of COVID-19 show the significant effects of the combined therapy of herbal medicine in treating COVID-19 without any significant side effects [135].Our results retrieved 91 in silico studies of natural products including herbal medicine. The results of this in silico approach showed that some of these studied active ingredients have a high affinity for each of the four important viral proteins compared with the inhibitors previously reported for each of these proteins. They could possibly have an inhibitory effect on the SARS-CoV-2 and COVID-19. Most evaluated chemical compounds had inhibitory effects against one or two proteins of SARS-CoV-2. In addition to having a great affinity to attach to the viral proteins, these herbal compounds have antioxidant, vasoprotective, anticarcinogenic, and antiviral properties. Thus, they can be applied as extremely safe therapeutic natural compounds and clinical assessments might have notable outcomes for controlling COVID-19.Due to the nature of preclinical (in vitro and in vivo) studies, the number of drug development studies in this area was less than in silico studies. Again, repurposed drugs were the most studied drugs in vitro (both herbal and synthetic). Some of them such as saquinavir, heparin, glycyrrhizic acid, and chlorpromazine show efficacy in both in vitro and in silico environments. Chlorpromazine is the single agent that was found to have efficacy in in silico, in vitro, and in vivo areas.In terms of mechanism of action, different targets from the structural and nonstructural proteins of COVID-19 were evaluated. Most of the studies focused on the main protease, papain-like protease, and spike glycoprotein. Apart from the specific protein that leads to viral replication, SARS-CoV-2 causes a surge of pro-inflammatory cytokines and chemokines, which cause damage to lung tissue and deterioration of lung function. Therefore, the design of a drug with multitarget of action against different proteins of COVID-19 and also anti-inflammatory potential could be valuable.Currently, there is no highly efficacious and specific treatment for SARS-CoV-2. Herein, we provided data on novel compounds in therapeutic drug discovery and development. Due to the nature of SARS-CoV-2 and the rises of several high transmissible strains, repurposing existing drugs has demonstrated power by bringing several drugs to approval for treating COVID-19 patients, such as remdesivir. Our results confirmed that a large number of repurposed agents are currently being explored for treating SARS-CoV-2 infection. However, these drugs still suffer from suboptimal therapeutic effect or known strong side effect. To accelerate drug discovery and development, especially during the current pandemic, natural products capture attention again. Our results showed that various natural bioactive compounds are being investigated in the preclinical step of drug development for COVID-19. In addition to having high affinity, these herb active ingredients have antioxidant, vasoprotective, anticarcinogenic, and antiviral properties. Therefore, they can be used as extremely safe therapeutic compounds in drug design studies to control COVID-19. However, the pharmacological effects and adverse reactions of some drugs under development are still unclear, and hence, well-designed high-quality studies are needed to further study the effectiveness and safety of these potential drugs to accelerate drug development targeting SARS-CoV-2 and thus promote progress towards ending the pandemic.



---

*Source: 2044282-2022-09-25.xml*