key: cord-1018254-4eokewei authors: Zhou, Qiongqiong Angela; Kato-Weinstein, Junko; Li, Yingzhu; Deng, Yi; Granet, Roger; Garner, Linda; Liu, Cynthia; Polshakov, Dmitrii; Gessner, Chris; Watkins, Steven title: Potential Therapeutic Agents and Associated Bioassay Data for COVID-19 and Related Human Coronavirus Infections date: 2020-07-30 journal: ACS Pharmacol Transl Sci DOI: 10.1021/acsptsci.0c00074 sha: 9f9327bbb55955d210c4a2e84de8ce5e6bc55312 doc_id: 1018254 cord_uid: 4eokewei [Image: see text] The COVID-19 pandemic, caused by the novel coronavirus SARS-CoV-2, has led to several million confirmed cases and hundreds of thousands of deaths worldwide. To support the ongoing research and development of COVID-19 therapeutics, this report provides an overview of protein targets and corresponding potential drug candidates with bioassay and structure–activity relationship data found in the scientific literature and patents for COVID-19 or related virus infections. Highlighted are several sets of small molecules and biologics that act on specific targets, including 3CLpro, PLpro, RdRp, S-protein–ACE2 interaction, helicase/NTPase, TMPRSS2, and furin, which are involved in the viral life cycle or in other aspects of the disease pathophysiology. We hope this report will be valuable to the ongoing drug repurposing efforts and the discovery of new therapeutics with the potential for treating COVID-19. COVID-19, the infectious disease caused by the newly emerged human coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), 1 was declared a global pandemic by the World Health Organization (WHO) on March 11, 2020. As of June 24, 2020, there have been more than 9 million confirmed cases and over 475,000 deaths worldwide. 2 In order to combat this pandemic and prevent future recurrences, scientists around the world have been working tirelessly to elucidate the molecular basis for SARS-CoV-2 infection and to develop effective therapeutic agents and preventative vaccines. SARS-CoV-2, a member of the Betacoronavirus genus, is an enveloped virus containing a single-stranded, positive-sense RNA genome. 3 Two other members of this genus that also cause similar severe acute respiratory diseases in humans are severe acute respiratory syndrome coronavirus (SARS-CoV or SARS-CoV-1) and Middle East respiratory syndrome coronavirus (MERS-CoV). As RNA viruses, they use their RNA-dependent RNA polymerase (RdRp) to replicate their genomic RNA. 4 In particular, SARS-CoV-2 and SARS-CoV share high levels of sequence homology and protein structural similarities. 5 They both use cell membrane protein angiotensin-converting enzyme 2 (ACE2) as their receptor and need host serine proteinase TMPRSS2 to cleave or "prime" their spike (S) protein in order to fuse with the host cell membrane and enter the cells. 6, 7 Once inside the host cells, the viral genome is translated by the host cell protein synthesis machinery and the resulting polyproteins are autoproteolytically cleaved by coronavirus proteases 3chymotrypsin-like protease (3CLpro) and papain-like protease (PLpro) to release smaller functional proteins to continue the viral replication process. 8−11 In some cases, an excessive immune response called a cytokine storm may contribute to the further development of pulmonary edema, acute respiratory distress syndrome (ARSD), and systemic inflammation. Mean-while, evidence is mounting that multiple organs may be damaged by SARS-CoV-2 infection in severe cases and that excessive blood coagulation may lead to life-threatening conditions. 12−14 While there is no specific treatment for COVID-19, over the past few months, significant advances in discovering the molecular mechanisms of SARS-CoV-2 infection have been made, and numerous clinical trials have begun, with many more in the planning stages. For instance, several antiviral drugs or drug candidates already approved for other diseases, such as lopinavir/ritonavir and darunavir (anti-HIV), remdesivir (Ebola), chloroquine and its derivatives (antimalarial), as well as Arbidol and favipiravir (broad spectrum antiviral), were among the first drugs to be tested in multiple clinical trials across the world. 15, 16 Camostat mesilate and nafamostat, both TMPRSS2 inhibitors, were enlisted in clinical trials shortly after the indispensable function of TMPRSS2 in SARS-CoV-2 infection was discovered. 7, 17 Since the uncontrolled inflammatory response is one of the major contributing factors to disease severity, anti-inflammatory drugs, such as interferon β, baricitinib, tocilizumab, sarilumab, and acalabrutinib, are also being evaluated in clinical trials for usage either alone or in conjunction with another anti-SARS-CoV-2 agent. 18−21 More recently, the potent anti-inflammatory effects of corticosteroids are being explored alone and/or in conjunction with other drugs. Initial reports showed that in patients hospitalized with COVID-19 dexamethasone resulted in a lower 28 day mortality among patients receiving respiratory support but not among those without respiratory support. 22 In addition, several clinical trials are either underway or being planned that look at the effects of dexamethasone alone or in combination with other drugs. 23 Although there are numerous ongoing clinical trials, only two drugs, remdesivir and favipiravir (avifavir), have so far been conditionally approved in a few countries for limited use, 24−26 and these appear to show only modest effects. Moreover, the application of remdesivir is further limited because it can only be administered intravenously to hospitalized patients. Additionally, despite some conflicting results, multiple studies and metaanalyses have concluded that hydroxychloroquine offers either very small or no therapeutic effects in the treatment of COVID-19. 27−29 The U.S. Food and Drug Administration (FDA) on June 15, 2020 revoked its Emergency Use Authorization for the use of hydroxychloroquine and chloroquine to treat COVID-19 30 and issued cautions against its use due to the risk of incurring heart rhythm problems and other safety issues, including blood and lymph system disorders, kidney injuries, and liver damage and failure. 31 However, clinical trials, including those on its use in prophylaxis, are continuing. 29 As a result, there is still an urgent need to identify effective therapeutic agents for COVID-19 and possible future coronavirus-related diseases. To support these efforts, we performed an analysis of the published journal articles and patents related to COVID-19 therapeutics. Herein, we review important viral and human targets and highlight target-based drug candidates with bioassay and structure−activity relationship (SAR) data from the Chemical Abstract Services (CAS)-indexed journal articles and patents. 2.1. Journal Analysis. Since the beginning of the COVID-19 outbreak, the number of journal articles published on this topic has continued to increase as shown in Figure 1 . Over the past 5 months, more than 16 000 articles have been published. This trend reflects the tremendous interest in the scientific community in understanding the new virus and in finding methods to combat the pandemic. A large number of publications are related to drug targets and the development of therapeutic agents. Due to the time requirements for de novo drug discovery, most efforts so far have focused on the repurposing of approved drugs, evaluation of investigational drugs (i.e., drugs in clinical trials for other purposes), molecular docking, and virtual screening studies. Table 1 highlights some notable journal articles, published during this period, which focused on the development and identification of therapeutics against COVID-19. These were selected based on a number of factors, including journal impact factor, the number of citations/downloads, and the type of studies. Because our focus is on newly studied drug candidates, articles on well-known drugs such as remdesvir, hydroxychloroquine, and others that were presented in our earlier report are not included here. 32 As shown in the table, both small molecules and biologics have been explored for the identification of COVID-19 therapeutics. 2.2. Patent Analysis. We also analyzed over 100 COVID-19-associated patents published in the first five months of 2020. These are categorized as follows: about 14% development of therapeutics including small molecules and biologics, 4% vaccines, 9% traditional Chinese medicines, and 56% the development of diagnostics. Table 2 lists specific patent applications related to COVID-19 therapeutics. Patent application CN111135167A discloses that GC376 (CAS Registry Number (RN) 1416992−39−6), a 3CLpro inhibitor, significantly reduces SARS-CoV-2 replication in cells with an EC 50 of 3.133 μM. Patent application CN111135166A discloses a pharmaceutical composition, consisting of GC376 and a prodrug, GS-441524 (CAS RN 1191237−69−0), which has a synergistic effect for inhibiting SARS-CoV-2 replication in cells with an EC 50 of 1.0 μM. Due to the different examination processes at various patent offices, it is likely that a significant number of COVID-19 therapeutics related patents will be published later this year. accessory proteins. The 16 NSPs are released by autoproteolysis of two large polyproteins by viral proteases, 3CLpro/NSP5 and PLpro/NSP3. 5 Table 3 summarizes the functions of the SARS-CoV-2 proteins as well as their sequence similarities with those from SARS-CoV. The proteins are grouped based on their functions: (1) NSPs related to viral proteolysis, (2) NSPs related to viral RNA modification or polymerization, (3) structural proteins involved in viral particle assembly, and (4) accessory proteins with various functions. As indicated in the table, most of the SARS-CoV-2 proteins share high sequence similarities with those of SARS-CoV. So far, most attention has been focused on Small-interfering nucleic acid, and its application for preparing pharmaceutical composition for preventing and/or treating new coronavirus pneumonia biologics CN111139241A 119 SARS-CoV-2 Small interfering nucleic acid for inhibiting new coronavirus and its composition and application biologics KR2020032050 120 SARS-CoV-2 COVID-19 virus customized triple knockout DNA treatment biologics the S protein, 3CLpro/NSP5, PLpro/NSP3, and RdRp/NSP12 as potential drug targets. These proteins not only serve crucial functions in the viral lifecycle of SARS-CoV-2 but also have been well-studied in the related viruses SARS-CoV and MERS-CoV. Although less-studied, other proteins, such as NSP7/8/9/10/ 13/14/15/16, may also serve as drug targets. Conceivably, those that interfere with host immune regulation (e.g., NSP1 and Orf3b/6/9b) may also be potential targets for anticytokine storm drugs. 5 3.2. Human Proteins Involved in SARS-CoV-2 Infection. Similar to other viruses, SARS-CoV-2 not only relies on its own proteins but also utilizes many proteins from host cells to achieve its attack on the host cells. These host proteins may also be potential drug targets, since they play crucial roles in one or more aspects of the disease, as shown in Tables 4 and S1. The proteins in Table 4 are grouped into very broad categories such as viral entrance, viral RNA/protein synthesis, host inflammatory response, and other functions. As an example of the human proteins involved in virus entrance, ACE2 functions as the main receptor for the S protein of SARS-CoV-2, 7 although other membrane proteins such as CD147/basigin may also be involved. 39, 40 After binding to a host cell receptor, the S protein needs to be cleaved by human proteases such as TMPRSS2, furin, or endosomal cathepsin L (CTSL) to initiate membrane fusion. 41 Additional host proteins involved in various steps of SARS-CoV-2 infection and abnormal host responses such as cytokine storm-mediated inflammation and excessive blood clotting 42 are also listed in Tables 4 and S1 . Clinical trial data was obtained as of 5/21/2020 from www.ClinicalTrials.gov. "Yes" in the table includes trials with the following status: "Not Yet Recruiting", "Recruiting", "Enrolling", "Active", or "Completed". Of the 20,000 journal articles and 2,200 patents found in our analysis, over 500 patents and more than 500 journal articles were identified that contain potential therapeutic substances against SARS-CoV, MERS-CoV, and SARS-CoV-2 infections. The associations of potential therapeutic substances with specific targets in these documents were determined by data mining of CAS-provided index entries followed by intellectual review of the results. Figure 2 shows some high-frequency document−potential therapeutic substance−protein target relationships. The protein targets are listed according to the number of documents associated with each target. As shown in the figure, the 20 targets include the structural proteins (S and N proteins), nonstructural proteins (3CLpro, RdRp, PLpro, and helicase/NTPase), and human host proteins (ACE2, DPP4, TMPRSS2, and furin). Most of these studies appeared to have focused on the identification and development of small molecule therapeutics, but some biologics (biosequences) have also been developed, including some targeting the S and N proteins. Detailed information about some selected anti-SARS-CoV-2, SARS-CoV, and MERS-CoV substances will be discussed in the subsequent section. 1. Data Sources. In order to identify drug candidates for COVID-19, we extracted SARS-CoV-2-associated bioassay data related to the development of therapeutics from recently published journals. We also examined bioassay data related to human coronaviruses published in journals and patents from 2000 to 2019, which contain substance information, targets, activity measures [half maximal inhibitory concentration (IC 50 ), half maximal effective concentration (EC 50 ), inhibition constant (K i ), and dissociation constant (K d )], and assay details. In this section, we focus on five viral proteins, 3CLpro, PLpro, RdRp, helicase/NTPase, and S protein, and two human proteases, TMPRSS2 and furin, that play a key role in S-protein-mediated cell entry of the virus. Selected substances with bioassay information toward these targets are presented in Tables 5−11 and in the Supporting Information. A high level view of the numbers of these substances associated with each protein target is found in Figure 3 . Of all the SARS-CoV-2 proteins, 3CLpro has the richest history of research data from other coronaviruses. Since 3CLpro is highly conserved among SARS-CoV-2, SARS-CoV, MERS-CoV, and other coronaviruses, previous research on this enzyme can serve as an excellent foundation for drug design of inhibitors of SARS-CoV-2 3CLpro. Table 5 highlights some substances that are active against 3CLpro of SARS-CoV-2 or SARS-CoV. Compounds GC376 and GC373 were designed based on the structures of 3CLpro from other viruses, but these were later shown to be also effective against SARS-CoV-2. 57, 58 Compounds 11a, 11b, 13a, and 13b were designed based on the recently revealed SARS-CoV-2 3CLpro crystal structure. 59, 60 In particular, 13a and 13b displayed longer plasma half-lives, and 13b can be nebulized for potential inhalant formulation. 59 As can be seen in the table, all these compounds share a common pyrrolidinyl structure. Some substances in Table 5 were initially identified in computer-based predictive modeling studies, which have greatly expedited the identification of potential 3CLpro inhibitors. For example, Li et al. performed molecular docking 61 Shown in Table 5 are three such drugs with relatively low IC 50 values, including dipyridamole, an anticoagulant currently in COVID-19 clinical trial, 62 and atazanavir, an HIV protease inhibitor with both anti-3CLpro and anti-inflammation activities. 63 Other clinically available protease inhibitors for other viruses, such as nelfinavir, 64 boceprevir, 57 and danoprevir, 65 were also found to be potent inhibitors of 3CLpro. In particular, danoprevir boosted by ritonavir showed promising results in COVID-19 patients. 65 Ebselen, an investigational drug with anti-inflammatory, antioxidant, and cytoprotective activities, has also been identified as 3CLpro inhibitor for SAS-CoV-2. 66, 67 Other drug candidates that functioned as cysteine protease inhibitors and inhibited SARS-CoV-2 infection include MDL-28170 and Z LVG CHN2, as identified from a large-scale drug repositioning screening of 12 000 FDA-approved and investigational drugs. 15 Carmofur, an antineoplastic drug, covalently binds to 3CLpro Cys145 (a critical residue in the catalytic site) and inhibits viral replication in Vero E6 cells. 68 3CLpro inhibitors discovered from SARS-CoV and MERS-CoV studies were also examined. A few of these compounds are shown in Table 5 , and a more complete list is given in Table S2 . Of these, both betulinic acid and savinin not only are 3CLpro inhibitors of SARS-CoV but also may act on other targets, with betulinic acid acting as a cannabinoid receptor (CB) modulator (CB1 antagonist/CB2 agonist) and savinin acting as a tumor necrosis factor α (TNF-α) antagonist. 69−72 Activation of cannabinoid receptor 2 (CB2), mainly expressed in immune cells, is reportedly linked to inhibition of inflammation and cytokine storms. 73, 74 Thus, activation of CB2 and inhibition of TNF-α would lead to attenuation of cytokine storm commonly observed in severe cases of COVID-19. While seemingly attractive as potential drug candidates for COVID-19, the polypharmacological properties of betulinic acid (inhibition of 3CLpro and activation of CB2) and savinin (inhibition of both 3CLpro and TNFα) remain to be confirmed. 75 In addition, several oligopeptides with or without chemical modification have been identified as 3CLpro inhibitors. For example, the octapeptide AVLQSGFR inhibited SARS-CoV 3CLpro and yet exhibited no cytotoxicity in Vero cells, indicating its potential as a drug candidate with low toxicity. 76, 77 5.3. Small-Molecule Inhibitors of PLpro. Besides its protease activity essential for viral replication, PLpro has the additional function of stripping ubiquitin and ISG15 from hostcell proteins to aid coronaviruses in escaping the host innate immune response. Therefore, inhibiting PLpro may be of use in not only inhibiting viral replication but also preventing the inhibition of innate immunity. 10 Table 6 presents selected small molecules shown to inhibit PLpro from SARS-CoV-2 or SARS-CoV. In a study with SARS-CoV-2 PLpro, two clinically safe Zn 2+ ejectors, disulfiram and ebselen 66 (also inhibit 3CLpro as shown in Table 5 ), were shown to extract Zn 2+ from the critical cysteine residues of PLpro and inhibit its enzyme activity. Tioguanine, also known as 6-thioguanine (6-TG), is a chemotherapy agent that is on the World Health Organization's List of Essential Medicines 81 and could potentially be used to treat COVID-19. A more complete list of substances active against PLpro can be found in Table S3 . Recently, the cryo-EM structure of RdRp has been revealed, 87 which will Table 7 provides compounds recently identified as inhibitors of SARS-CoV-2 RdRp in various bioassays. Included in this table are some FDA-approved drugs, such as sofosbuvir (a key component of hepatitis C drug EPCLUSA), azidothymidine (an anti-HIV drug), tenofovir alafenamide (a drug for HIV and hepatitis B), and tenofovir and emtricitabine (two components in DESCOVY and TRUVADA, two anti-HIV drugs). 88, 89 In addition, previously discovered SARS-CoV RdRp inhibitor EIDD-1931 was tested in SARS-CoV-2 and exhibited a high potency for infection inhibition. 91 Its oral form, EIDD-2801, was also tested in animal models. 91 A complete list of substances active against other (+)ssRNA viruses is shown in Table S4 . That Affect Viral Entry Mediated by S-Protein−ACE2 Interactions. Unlike the viral proteases and RdRp, which are more likely to be inhibited by small molecules, inhibitors of the interaction of S protein with receptor ACE2 are predominantly small peptides and recombinant proteins mimicking ACE2 or neutralizing antibodies against the S protein. Recently, many of these biological molecules have been tested with SARS-CoV-2, as shown in Table 8 . For example, when EK1, a peptidic pancoronavirus fusion inhibitor which targets the heptad repeat (HR)1 region of the S protein, was linked to cholesterol (EK1C and EK1C4) or palmitic acid (EK1P), they displayed more potent inhibition against SARS-CoV-2 S-protein-mediated membrane fusion. 92 Another lipopeptide, IPB02, is designed based on HR2 sequence and also showed strong activity in inhibiting the SARS-CoV-2 S-protein-mediated viral−cell fusion. 93 SBP1, derived from the α1 helix of ACE2 peptidase domain, showed high affinity to the SARS-CoV-2-RBD. 94 Recombinant proteins ACE2-Fc and hrsACE2, which act as decoy receptors, also target the S-protein−ACE2 interaction and viral−host-cell membrane fusion. 95, 96 Furthermore, an increasing number of antibodies, immunoglobulin fragments, or In addition to the inhibitors mentioned above that have been tested with SARS-CoV-2, we found more compounds from SARS-CoV experiments that could be valuable for SARS-CoV-2 treatment. For example, the small molecule VE607 inhibits both S-protein−ACE2 interaction-mediated SARS-CoV entry and SARS-CoV plaque formation. 123 The flavonoid luteolin has been reported to bind to the S protein and inhibit SARS-CoV entry into host cells. 124, 125 It also has anti-inflammatory and 3CLpro inhibition activities. 126 5.6. Small-Molecule Inhibitors of SARS-CoV Helicase/ NTPase. As mentioned earlier in this report, NSP13 displays both helicase and NTPase activities and initiates the first step in viral mRNA capping. As part of a complex with NSP14 and NSP16, it installs the cap structure onto viral RNA in the cytoplasm. Since the sequence of SARS-CoV-2 helicase/ NTPase is almost identical (100% in sequence similarity) to that of SARS-CoV, 129 inhibitors of SARS-CoV helicase/NTPase will most likely work for SARS-CoV-2 as well. Specific examples of such inhibitors are shown in Table 9 , and a more complete list is given in Table S5 . Some trioxaadamantanetriol compounds, such as bananin and vanillinbananin, inhibited replication of SARS-CoV in cultured cells with low cytotoxicity. 130, 131 In addition, the plant-derived flavonoids myricetin and scutellarein, which are both found in tea, have been shown as active inhibitors with low toxicity. 132, 133 Unlike the above mentioned inhibitors, SSYA10−001 demonstrated helicase inhibition without affecting the cellular ATPase activity. 134 TMPRSS2. Human serine protease TMPRSS2 is involved in S protein priming needed for the S2 segment of the S protein to mediate fusion of the viral envelope with the host cell membrane. 7 Selected inhibitors are shown in Table 10 , and a more complete list is given in Table S6 . In addition to their inhibitory effect on TMPRSS2, these selected inhibitors are known to have other functions that may be beneficial in treating COVID-19. For example, bicalutamide, enzalutamide, dimethylcurcin, and CAS RN 2031161−35−8 are nonsteroidal antiandrogen drugs that were shown to inhibit TMPRSS2 expression. 137 Since TMPRSS2 is an androgen-regulated gene that is overexpressed in prostate cancer, 138 speculation has arisen that higher androgen levels could be the reason for more severe outcomes in men with COVID-19. 139 In addition, inhibitors of androgen signaling have been shown to reduce ACE2 levels; therefore, these inhibitors may have dual functions affecting both ACE2 and TMPRSS2. 140 Finally, compounds MI460 and CAS RN 944925−37−5 may also inhibit proinflammatory cytokines and block blood coagulation-related factors, respectively. 141, 142 5.8. Small-Molecule and Peptide Inhibitors of Human Protease Furin. The human protease furin is a ubiquitously expressed subtilisin/kexin-like proprotein convertase (PC) that 145 Similar to TMPRSS2, furin is involved in priming viral S protein to mediate viral fusion with the host cell membrane and subsequent viral entry. Its cleavage site (RRAR↓) at the S1/S2 boundary of the SARS-CoV-2 and MERS-CoV S protein matches the minimal requirement of furin substrate sequence. 146, 147 Many furin inhibitors have been reported in the literature. Selected substances are shown in Table 11 , and a more complete list is given in There are also nonpeptidic furin inhibitors, such as the guanidinylated aryl 2,5-dideoxystreptamine-derived compound represented by CAS RN 922732−52−3. This substance inhibited not only furin but also other PC family members (PC6B, PACE4, and PC7) without significant cytotoxicity to cells. 150 Another inhibitor, oroxylin A, an O-methylated flavone natural product extracted from Scutellaria roots, has antiinflammatory and anticoagulation activities, which may also be beneficial in treating COVID-19. 151 Baicalein, a flavone from the roots of Scutellaria baicalensis, has been shown to inhibit Dengue virus replication in Vero cells and has also been reported recently to inhibit SARS-CoV-2 3CLpro. 152 In addition, baicalein has antibacterial and anti-inflammatory activities. 79 Although these inhibitors may be used to treat COVID-19 and its associated complications, more study is needed to ensure the safety of these compounds. 5.9. Small Molecules and Biologics Targeting Other Human Proteins Involved in SARS-CoV-2 Infection. In addition to TMPRSS2 and furin, there are many other human proteins as listed in Table 4 which have been shown to be involved in COVID-19. Table 12 lists several small molecules or biologics targeting these human proteins involved in different steps of SARS-CoV-2 infection. A number of these, including Table 9 . Small-Molecule Inhibitors of Helicase/NTPase 80, 123, 130, 133, 135, 136 a "*": multiple activity measure values for one substance are from multiple references. In light of the enormous amount of published information and rapidly evolving knowledge about COVID-19, this report systematically assembles and curates a large amount of data into one resource to support the ongoing research and development of COVID-19 therapeutics. Highlighted are notable journal articles and patents related to COVID-19, important viral and human protein targets, a high-level view of target−substance relationship in documents related to COVID-19, SARS, and MERS, as well as rich lists of target-based potential drug candidates for COVID-19 and related coronavirus infections. The potential drug candidates include both smalland large-molecule biologics. The small molecules are comprised of a wide variety of organic compounds, nucleotide analogs, and peptides, while the biologics are mainly antibodies along with a few recombinant proteins. More importantly, we report bioassay data with detailed structure−activity relationship information extracted from published studies. We hope this report will be valuable to the ongoing drug repurposing efforts and the discovery of new therapeutics with the potential for treating COVID-19. It is worth mentioning that although these preclinical studies provide important information the utility of the listed substances as drugs for COVID-19 or related coronavirus infections would ultimately rely on successful clinical trials. In addition to the various wet-laboratory-based approaches, computational drug repurposing for COVID-19 also plays a significant role in accelerating therapeutic development for this and other diseases. In this approach, a variety of computational and clinical data are often used and analyzed together for drug repurposing. 165 This approach can help to overcome the challenge of translating basic scientific findings to human applications, because these drugs have passed clinical safety and bioavailability testing, thereby increasing their chances for final approval. 166 For example, in a high-throughput docking approach, after screening a chemical library built from FDAapproved drugs and compounds undergoing clinical trials, Cavasotto and Di Filippo identified several structurally diverse compounds that each displayed antiviral activity against SARS-CoV-2. 167 Another structure-based virtual study suggested that toremifene, an FDA-approved estrogen receptor modulator for treating advanced breast cancer, may inhibit the SARS-CoV-2 S protein and methyltransferase/NSP14. 168 Moreover, melatonin was identified by the network medicine approach as showing a 171 It can be expected that computer-screening of compounds and modeling will be increasingly used in the discovery of drugs for COVID-19 and other viral infections to expedite the drug development process and lower its cost. Nevertheless, experimental evaluation of drug candidates' efficacy in cell-based assays and animal model studies is still needed to confirm the suggested drug effects of these virtually selected molecules. Although this paper focuses on individual therapeutics, therapy regimes that combine various drugs to target multiple pathological processes and/or molecular targets have been evaluated and may play an important role in treating COVID-19. Over 600 documents covered drug combination approaches for COVID-19 in the CAS scientific literature collection. These include studies on the well-publicized hydroxychloroquine and azithromycin combination and on remdesivir with numerous other drugs. Of the latter, one interesting paper that combined in silico and in vitro methods highlighted the combination of remdesivir with nitazoxanide. 172 Since COVID-19 is often characterized by exaggerated inflammatory responses, anti- Since COVID-19 patients with an underlying condition, such as cardiovascular disease or diabetes, are more likely to be hospitalized and have life-threatening conditions, it is very likely that COVID-19 patients receiving antiviral drugs are simultaneously on other medications for their pre-existing conditions. Therefore, it is also crucial that COVID-19 drugs given should be compatible with those medications that the patient is already taking in order to prevent undesirable drug−drug interactions. Currently, it is unknown how long the COVID-19 crisis will last. As different parts of the world become increasingly interconnected, it seems likely that there will be additional pandemics in the years to come, and many of these will be of viral origin. We hope the current focus on antiviral agent research will lead to major breakthroughs and help us to be better prepared for future outbreaks. The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsptsci.0c00074. Table S1 : a more complete list of human protein targets involved in COVID-19, Table S2 : inhibitors of 3CLpro in SARS-CoV and MERS-CoV, A Novel Coronavirus from Patients with Pneumonia in China Genomic characterization and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding Emergence of a Novel Coronavirus, Severe Acute Respiratory Coronavirus 2: Biology and Therapeutic Options A SARS-CoV-2 protein interaction map reveals targets for drug repurposing Structural basis of receptor recognition by SARS-CoV-2 SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 is Blocked by a Clinically Proven Protease Inhibitor Therapeutic options for the 2019 novel coronavirus (2019-nCoV) Learning from the Past: Possible Urgent Prevention and Treatment Options for Severe Acute Respiratory Infections Caused by 2019-nCoV The SARS-coronavirus papain-like protease: Structure, function and inhibition by designed antiviral compounds Genomic and protein structure modelling analysis depicts the origin and infectivity of 2019-nCoV The Trinity of COVID-19: immunity, inflammation and intervention Cytokine release syndrome in severe COVID-19 COVID-19 and Thrombotic or Thromboembolic Disease: Implications for Prevention, Antithrombotic Therapy, and Follow-Up Discovery of SARS-CoV-2 antiviral drugs through large-scale compound repurposing The anti-influenza virus drug, Arbidol is an efficient inhibitor of SARS-CoV-2 in vitro Identification of nafamostat as a potent inhibitor of Middle East respiratory syndrome coronavirus Smediated membrane fusion using the split-protein-based cell-cell fusion assay Effect of Treatments in Patients Hospitalized for Severe COVID-19 Pneumonia: A Multicenter Cohort Study Inhibition of Bruton tyrosine kinase in patients with severe COVID-19 Dexamethasone in Hospitalized Patients with Covid-19 -Preliminary Report Fact Sheet for Health Care Providers Emergency Use Authorization (EUA) of Remdesivir (GS-5734) Glenmark Initiates Phase 3 Clinical Trials on Antiviral Favipiravir for COVID-10 Patients in India. Glenmark Pharmaceuticals Ltd., News release Russian Ministry of Health approves the first COVID-19 drug Avifavir produced by JV of RDIF and ChemRar. Russian Direct Investment Fund News release Observational Study of Hydroxychloroquine in Hospitalized Patients with Covid-19 (30) U.S. Food & Drug Administration. (2020) Revoke of EUA for emergency use of oral formulations of chloroquine phosphate (CQ) and hydroxychloroquine sulfate (HCQ FDA cautions against use of hydroxychloroquine or chloroquine for COVID-19 outside of the hospital setting or a clinical trial due to risk of heart rhythm problems Research and Development on Therapeutic Agents and Vaccines for COVID-19 and Related Human Coronavirus Diseases Ultrastructure and Origin of Membrane Vesicles Associated with the Severe Acute Respiratory Syndrome Coronavirus Replication Complex RNA 3′-end mismatch excision by the severe acute respiratory syndrome coronavirus nonstructural protein nsp10/nsp14 exoribonuclease complex The Contribution of the Cytoplasmic Retrieval Signal of Severe Acute Respiratory Syndrome Coronavirus to Intracellular Accumulation of S Proteins and Incorporation of S Protein Into Virus-Like Particles Conserved Domain in the Coronavirus Membrane Protein Tail Is Important for Virus Assembly SARS-CoV ORF6 Antagonizes STAT1 Function by Sequestering Nuclear Import Factors on the rER/ Golgi membrane CD147 as a Target for COVID-19 Treatment: Suggested Effects of Azithromycin and Stem cell engagement SARS-CoV-2 invades host cells via a novel route: CD147-spike protein Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV Bioinformatic characterization of angiotensin-converting enzyme 2, the entry receptor for SARS-CoV-2. bioRxiv Mutations, recombination and insertion in the evolution of 2019-nCoV. bioRxiv Cell entry mechanisms of SARS-CoV-2 JAK1 inhibition blocks lethal sterile immune responses: implications for COVID-19 therapy. bioRxiv Baricitinib therapy in COVID-19: A pilot study on safety and clinical impact The anti-viral facet of anti-rheumatic drugs: Lessons from COVID-19 The first case of COVID-19 treated with the complement C3 inhibitor AMY Exuberant elevation of IP-10, MCP-3 and IL-1ra during SARS-CoV-2 2 infection is associated with disease severity and fatal outcome. medRxiv Neutrophil extracellular traps (NETs) as markers of disease severity in COVID-19. medRxiv The IMPDH inhibitor merimepodib suppresses SARS-CoV-2 replication in vitro Broad-spectrum antiviral activity of the eIF4A inhibitor silvestrol against corona-and picornaviruses Integrity of the Early Secretory Pathway promotes, but Is Not Required for, Severe Acute Respiratory Syndrome Coronavirus RNA Synthesis and Virus-Induced Remodeling of Endoplasmic Reticulum Membranes Proteomics of SARS-CoV-2-infected host cells reveals therapy targets Structural Genomics of SARS-CoV-2 Indicates Evolutionary Conserved Functional Regions of Viral Proteins Boceprevir, GC-376, and calpain inhibitors II, XII inhibit SARS-CoV-2 viral replication by targeting the viral main protease Feline coronavirus drug inhibits the main protease of SARS-CoV-2 and blocks virus replication Crystal structure of SARS-CoV-2 main protease provides a basis for design of improved αketoamide inhibitors Structure-based design of antiviral drug candidates targeting the SARS-CoV-2 main protease Identify potent SARS-CoV-2 main protease inhibitors via accelerated free energy perturbation-based virtual screening of existing drugs Therapeutic effects of dipyridamole on COVID-19 patients with coagulation dysfunction. medRxiv Atazanavir inhibits SARS-CoV-2 replication and proinflammatory cytokine production. bioRxiv First clinical study using HCV protease inhibitor to treat naive and experienced COVID-19 patients. medRxiv Multi-Targeting of Functional Cysteines in Multiple Conserved SARS-CoV-2 Domains by Clinically Safe Zn-ejectors Structure of M pro from COVID-19 virus and discovery of its inhibitors Structural basis for the inhibition of SARS-CoV-2 main protease by antineoplastic drug carmofur Specific Plant Terpenoids and Lignoids Possess Potent Antiviral Activities against Severe Acute Respiratory Syndrome Coronavirus Betulinic acid targets YY1 and ErbB2 through cannabinoid receptor-dependent disruption of microRNA-27a:ZBTB10 in breast cancer Small Molecules from Nature Targeting G-Protein Coupled Cannabinoid Receptors: Potential Leads for Drug Discovery and Development. Evidence-Based Complementary Altern Savinin, a lignan from Pterocarpus santalinus inhibits tumor necrosis factor-SSproduction and T cell proliferation Cannabinoid Receptor Type 2: A Possible Target in SARS-CoV-2 (CoV-19) Infection? Cannabinoids as novel anti-inflammatory drugs Trials of antitumour necrosis factor therapy for COVID-19 are urgently needed Discovery of Potent Anti-SARS-CoV MPro Inhibitors Polyprotein cleavage mechanism of SARS CoV Mpro and chemical modification of the octapeptide 2020) Scutellaria baicalensis extract and baicalein inhibit replication of SARS-CoV-2 and its 3C-like protease in vitro Discovery of baicalin and baicalein as novel, natural product inhibitors of SARS-CoV-2 3CL protease in vitro Antiviral drug discovery against SARS-CoV Thiopurine analogues inhibit papain-like protease of severe acute respiratory syndrome coronavirus Structure-Based Design, Synthesis, and Biological Evaluation of a Series of Novel and Reversible Inhibitors for the Severe Acute Respiratory Syndrome-Coronavirus Papain-Like Protease Severe Acute Respiratory Syndrome Coronavirus Papain-like Novel Protease Inhibitors: Design, Synthesis, Protein-Ligand X-ray Structure and Biological Evaluation A noncovalent class of papain-like protease/deubiquitinase inhibitors blocks SARS virus replication Compounds and methods for treating respiratory diseases, WO2010022355A1, World Intellectual Property Organization Structure of RNAdependent RNA polymerase from 2019-nCoV, a major antiviral drug target Nucleotide analogues as inhibitors of SARS-CoV-2 polymerase. bioRxiv A library of nucleotide analogues terminate RNA synthesis catalyzed by polymerases of coronaviruses causing SARS and COVID-19 An orally bioavailable broad-spectrum antiviral inhibits SARS-CoV-2 in human airway epithelial cell cultures and multiple coronaviruses in mice Design of Potent Membrane Fusion Inhibitors Against SARS-CoV-2, an Emerging Coronavirus With High Fusogenic Activity Investigation of ACE2 N-terminal fragment binding to SARS-CoV-2 Spike RBD. bioRxiv Potential host range of multiple SARS-like coronaviruses and an improved ACE2-Fc variant that is potent against both SARS-CoV-2 and SARS-CoV-1. bioRxiv Inhibition of SARS-CoV-2 Infections in Engineered Human Tissues Using Clinical-Grade Soluble Human ACE2 Immunoglobulin fragment F(ab′)2 against RBD potently neutralizes SARS-CoV-2 in vitro Potent Neutralizing Antibodies against SARS-CoV-2 Identified by High-Throughput Single-Cell Sequencing of Convalescent Patients' B Cells Humanized single domain antibodies neutralize SARS-CoV-2 by targeting spike receptor binding domain A human monoclonal antibody blocking SARS-CoV-2 infection Computational design of ACE2-based peptide inhibitors of SARS-CoV-2 COVID-19: combining antiviral and anti-inflammatory treatments COVID-19: immunopathology and its implications for therapy Development of CRIPSR as an Antiviral Strategy to Combat SARS-CoV-2 and Influenza Rapid identification of potential inhibitors of SARS-CoV-2 main protease by deep docking of 1.3 billion compounds Repurposing therapeutics for COVID-19: Supercomputerbased docking to the SARSCoV-2 viral spike protein and viral spike protein-human ACE2 interface Application of silver monoethyl fumarate in resisting novel coronavirus infection, CN111184708A, People's Republic of China Use of tolfenamic acid or a pharmaceutically acceptable salt thereof in the preparation of medicament for preventing and/or treating novel coronavirus inflammation, CN111184707A, People's Republic of China Application of LTX-315 in preparing products for inhibiting coronavirus, CN111150833A, People's Republic of China Application of GS-441524 in preparing novel coronavirus SARS-CoV-2 inhibitor, CN111135184A, People's Republic of China Application of GC376 in preparation of novel coronavirus SARS-CoV-2 inhibitor, CN111135167A, People's Republic of China Pharmaceutical composition consisting of GC376 and GS-441524 and application thereof in inhibiting novel coronavirus, CN111135166A, People's Republic of China Application of 2019-nCoV 3CL hydrolase inhibitor and IL-6 monoclonal antibody in preparing medicament or treating coronavirus disease 2019, CN111053909A, People's Republic of China Composition containing benzylisoquinoline alkaloid and trans-resveratrol for treating coronavirus infection, CN110960532A, People's Republic of China Overexpression ACE2 mesenchyma cell in the preparation of medicine for treating new coronavirus application of drugs and preparation method thereof, CN111166768A, People's Republic of China Preparation method of gene therapy product for treating COVID-19, CN111172195A, People's Republic of China A human SARS-CoV-2 monoclonal antibody and preparation method and application thereof, CN111153991A, People's Republic of China SARS-CoV-2 Small-interfering nucleic acid, and its application for preparing pharmaceutical composition for preventing and/or treating new coronavirus pneumonia, CN111139242A, People's Republic of China Small interfering nucleic acid for inhibiting new coronavirus and its composition and application, CN111139241A, People's Republic of China COVID-19 virus customized triple knockout DNA treatment Rapid selection of a human monoclonal antibody that potently neutralizes SARS-CoV-2 in two animal models Cross-neutralization of SARS-CoV-2 by a human monoclonal SARS-CoV antibody Identification of Novel Small-Molecule Inhibitors of Severe Acute Respiratory Syndrome-Associated Coronavirus by Chemical Genetics Small molecules blocking the entry of severe acute respiratory syndrome coronavirus into host cells Targeting SARS-CoV-2 Spike Protein of COVID-19 with Naturally Occurring Phytochemicals: An in Silco Study for Drug Development Inhibition of SARS-CoV 3CL protease by flavonoids Identification of a minimal peptide derived from heptad repeat (HR) 2 of spike protein of SARS-CoV and combination of HR1-derived peptides as fusion inhibitors Design and biological activities of novel inhibitory peptides for SARS-CoV spike protein and angiotensin-converting enzyme 2 interaction The Proteins of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS CoV 2 or n-COV19), the Cause of COVID 19 An update on the bananins: anti-RNA-viral agents with unique structural signature The Adamantane-Derived Bananins Are Potent Inhibitors of the Helicase Activities Profiles of Potentially Antiallergic Flavonoids in 27 Kinds of Health Tea and Green Tea Infusions Identification of myricetin and scutellarein as novel chemical inhibitors of the SARS coronavirus helicase, nsP13 Severe acute respiratory syndrome coronavirus replication inhibitor that interferes with the nucleic acid unwinding of the viral helicase Suppression of SARS replication by SARS helicase inhibitors, WO2013188887A1, World Intellectual Property Organization The bananins: new anticorona-RNA-viral agents with unique structural signature Select androgen receptor degrader (SARD) ligands and methods of use thereof, US20170029370A1 and US10017471B2, US Patent and Trademark Office The Androgen-Regulated Protease TMPRSS2 Activates a Proteolytic Cascade Involving Components of the Tor Microenvironment and Promotes Prostate Cancer Metastasis Sex hormones signal why virus hits men harder Androgen Regulates SARS-CoV Receptor Levels and Is Associated with Severe COVID-19 Symptoms in Men. bioRxiv The Impact of Acute Matriptase Inhibition in Hepatic Inflammatory Model Development of substrate analogue inhibitors for the human airway trypsin-like protease HAT Use of TMPRSS2 inhibitors as medicaments, WO2013014074A1, World Intellectual Property Organization Use of HAT inhibitors and TMPRSS2 inhibitors as medicaments, WO2010149459A1, World Intellectual Property Organization Elongated and Shortened Peptidomimetic Inhibitors of the Proprotein Convertase Furin Multibasic Cleavage Site in the Spike Protein of SARS-CoV-2 Is Essential for Infection of Human Lung Cells The spike glycoprotein of the new coronavirus 2019-nCoV contains a furin-like cleavage site absent in CoV of the same clade Therapeutic uses of furin and its inhibitors: a patent review Decanoyl-Arg-Val-Lys-Arg-Chloromethylketone: An Antiviral Compound That Acts against Flaviviruses through the Inhibition of Furin-Mediated prM Cleavage Synthetic small molecule furin inhibitors derived from 2,5-dideoxystreptamine Overview of Oroxylin A: A Promising Flavonoid Compound Baicalein: A review of its anti-cancer effects and mechanisms in Hepatocellular Carcinoma Inhibition of tumor cells proliferation and migration by the flavonoid furin inhibitor isolated from Oroxylum indicum Proprotein convertase inhibitory activities of flavonoids isolated from Oroxylum indicum Modeling the activity of furin inhibitors using artificial neural network New substrate analogue furin inhibitors derived from 4-amidinobenzylamide Cleavage targets and the D-arginine-based inhibitors of the West Nile virus NS3 processing proteinase Targeting host proteinases as a therapeutic strategy against viral and bacterial pathogens Teicoplanin potently blocks the cell entry of 2019-nCoV. bioRxiv The inhaled corticosteroid ciclesonide blocks coronavirus RNA replication by targeting viral NSP15. bioRxiv Inhibition of PIKfyve kinase prevents infection by Zaire ebolavirus and SARS-CoV-2. bioRxiv Multidrug treatment with nelfinavir and cepharanthine against COVID-19. bioRxiv Pulmonary radiological change of COVID-19 patients with 99mTc-MDP treatment. medRxiv Consideration of dornase alfa for the treatment of severe COVID-19 acute respiratory distress syndrome Drug repurposing: progress, challenges and recommendations Lost in translation: the valley of death across preclinical and clinical divide -identification of problems and overcoming obstacles silico Drug Repurposing for COVID-19: Targeting SARS-CoV-2 Proteins through Docking and Quantum Mechanical Scoring. ChemRxiv Repurposing of FDA-Approved Toremifene to Treat COVID-19 by Blocking the Spike Glycoprotein and NSP14 of SARS-CoV-2 A Network Medicine Approach to Investigation and Population-based Validation of Disease Manifestations and Drug Repurposing for COVID-19 Repurpose Open Data to Discover Therapeutics for COVID-19 using Deep Learning Drug repurposing for coronavirus (COVID-19): in silico screening of known drugs against coronavirus 3CL hydrolase and protease enzymes Discovery of Synergistic and Antagonistic Drug Combinations against SARS-CoV-2 2020) Network-based drug repurposing for novel coronavirus 2019-nCoV/SARS-CoV-2 ACS Pharmacology & Translational Science pubs.acs.org/ptsci Review