**4. Repurposing therapeutics towards SARS-CoV-2 potential targets**

Therapeutics facilities aid physicians and scientist to help patients with the better cure for the disease and develop a vaccine. The production of novel therapeutics and vaccination is now at the earliest possible time to curb the epidemic. In addition, continuous attempts are being made at regional, national, and person levels to recognize the host, genomics, epidemiological interactions, and propagation modes of SARS-CoV-2 in order to control the epidemic. Scientists worldwide are actively working to establish an optimal antiviral agent and vaccine against SARS-CoV-2 [51]. One way of preventing this issue is to test two or three drugs with minimal prolixity that operate on various cellular signals with viral replication. A further approach that helps researchers to curb the variety of individual antimicrobials for emerging and re-emerging infectious diseases is the high-performance screening of host-virus interaction level composite libraries for synergistic combinations [52, 53]. Nevertheless, the screening of experimental small molecule drugs in combinations and other potent anti-SARS-CoV-22 agents with new and improved key characteristics can be effective which can be helpful in COVID-19 therapy. Potential medicines include previously used or tested medicines for diagnosing diseases and medicines recently discovered or developed. In most cases, the benefits of drug repurposing are safety evaluation, preclinical monitoring, and in some cases the development processes would not take much time and thus reduce the time needed for drug development. The chances of failure with repurposed drugs would be smaller as the early steps of medication efficacy and safety testing have already been completed and the treatment has already been shown to be sufficiently effective in a preclinical and human model [54]. Antiviral therapies are being investigated towards treatment of COVID-19 because the SARS-CoV-2 replication of leads to wide variation of the clinical manifestations. As significant functional proteins of SARS-CoV-2, Nsps and other structural and accessory proteins are involved in transcription of RNA, synthesis of protein, post-translational modification, viral duplication and contagion of the host. Among them, PLpro (nsp3), 3CLpro (nsp5), RdRp (nsp12), Helicase (nsp13), Spike glycoprotein (S) and Nucleocapsid (N) are the most important targets for the development of small-molecule inhibitors due to the enzyme active site and clear biological functions [55].

#### **4.1 Protease inhibitors**

Protease inhibitors are drugs that inhibit the activity of protease enzyme responsible for viral development, infection, and replication by cleaving it into smaller fragments. They bind to the active site of the enzyme, mediate and block the maturation of freshly formed virions [51]. Two major targets: PLpro and 3-CLpro in the replicase 1a and 1ab domain are essential component for virus reproduction and regulation of host cell response thus being the important target for the SARS-CoV-2 inhibitors (**Figure 9**). Ribavirin (SCH-18908) [NCT00578825], an antiviral drug acts on the replicase protein, preventing binding of the nucleotides, resulting in reduced viral replication or the formation of defective virions. Lopinavir [NCT04321174] and Ritonavir [NCT04330690], are protease inhibitors used to treat HIV infections by inhibiting the HIV protease enzyme to form an inhibitor-enzyme complex and proteolytic cleavage of the viral polyprotein precursors [3]. Darunavir [NCT01448707], another (HIV-1) nonpeptidic protease inhibitor with the inhibition activity of the dimerization and catalytic activity of HIV-1 protease. Saquinavir [DB01232] inhibiting HIV1/2 protease-mediated lysis of HIV gag and pol polyproteins was found cytotoxically active at conc. Above 50 μM [56]. Rupintrivir (DB05102) a broad-spectrum

**Figure 9.** *Repurposed protease inhibitors for COVID-19 treatment.*

antiviral agent is a potent 3C-Like protease inhibitor against norovirus, picornavirus, and coronavirus proteases in development against human rhinoviral (HRV) infections that has been recently co-crystallized with its target [57]. Larazotide acetate (AT1001) (NCT03569007) a paracellular permeability peptide inhibitor has been studied for the treatment of autoimmune diseases such as type I diabetes mellitus, gastrointestinal diseases and disorders. Studies have revealed that AT1001 potentially binds to the Mpro catalytic domain showing up intermolecular interactions [58]. Teicoplanin (DB06149), a glycopeptide antibiotic widely used to treat bacterial infections, has been reported to be active against SARS-CoV-2 in *in-vitro* studies inhibiting peptidoglycan polymerization, resulting in inhibition of bacterial cell wall synthesis and cell death [59, 60]. Nafamostat mesylate [NCT04418128], a clinical proven and synthetic serine protease inhibitor has been identified to inhibit the activity SARS-CoV-2 by targeting TMPRSS2-dependent host cell entry [61, 62]. However, *in-vitro* studies of active phytochemical compounds from the natural sources have revealed the SARS-CoV-2 proteolytic and inhibitory activity; Phaitanthrin D (*Isatis indigotica Fort*), Baicalin (*Scutellaria baicalensis*), Piceatannol (*Vitis vinifera*), Platycodin D (*Platycodon grandiflorus*), Betulonal (*Cassine xylocarpa*), 2b-Hydroxy-3,4-secofriedelolactone-27-oic acid (*Viola diffusa*), Cleistocaltone A (*Cleistocalyx operculatus*), Phyllaemblinol (*Phyllanthus emblica*) etc. [55].

### **4.2 Replicase inhibitors**

RdRp is a critical non-structural protein component of the SARS-CoV-2 genome that uses a metal-ion-dependent mechanism to catalyze viral RNA synthesis. However, due to its comprehensive knowledge about the different domains and its functions, strong preservation among evolutionary RNA viruses and the lack

#### *Repurposed Therapeutic Strategies towards COVID-19 Potential Targets Based on Genomics… DOI: http://dx.doi.org/10.5772/intechopen.96728*

of homologous sequences in mammalian cells; the ease progress and consequent access to biochemical assays to quickly detect large collections of compounds [63]. Remdesivir (GS-5734) [NCT04252664], a nucleoside analogue acts on the replicase polyprotein 1ab of SARS-CoV-2 genome preventing RNA polymerase function resulting in ending the transcription of RNA and reduces the production of viral RNA. Penciclovir [NCT00820534], another nucleoside analogue, synthetic acyclic guanine derivative with antiviral activity targets the RdRp inhibiting the DNA synthesis of virus-infected cells and terminating viral replication (**Figure 10**). β-D-N4-hydroxycytidine, an orally bioavailable, broad-spectrum antiviral ribonucleoside analogous to multiple RNA viruses like influenza, CoV, equine encephalitis and Ebola viruses have been found in *in-vitro* studies of antiviral effect on SARS-CoV-2 primarily through mutagenesis of viral RNA [64, 65]. Cefuroxime, an anti-bacterial drug is prescribed in patients with COVID-19, as a potential inhibitor with the ability of binding tightly to the active site of the enzyme, with a highest ICM score of −41.30, and mfscore of −63.04, which when compared to Remdesivir had a score of −27.4 and a mfscore of −113 [66] The natural products and derivatives with antivirus, anti-inflammation and anti-tumor effects exhibited high binding affinity to RdRp, such as 14-deoxy-11,12-didehydroandrographolide (*Andrographis paniculate*), Gnidicin (*Gnidia lamprantha*), 2b,30b-dihydroxy-3,4-seco-friedelolactone-27-lactone (*V. diffusa*), Theaflavin 3,30-di-O-gallate (*Camellia sinensis*), Betulonal (*C. xylocarpa*), 1,7-dihydroxy-3- methoxyxanthone (*Swerti apseudochinensis*) [67].

SARS-CoV-2 replication enzyme, helicase has the properties of unwinding and splitting DNA and RNA into two single-stranded nucleic acids and these

**Figure 10.**

*Repurposed replicase inhibitors for COVID-19 treatment.*

characteristic of the enzyme marks it as a potential target to be studied [68]. Bananins have been shown to inhibit SARS-CoV ATPase activity leading to inhibition of viral replication *in-vitro* with IC50 values far less than 10 mM [69] Bismuth salt, such as Bismuth potassium citrate (BPC) has been observed to dosedependently inhibit both the NTPase and RNA helicase activities of SARS-CoV-2 nsp13. Further studies indicates that SARS-CoV-2 nsp13 may be a valuable target for antivirals, which could be beneficial in attempting to regulatory mechanism of this life-threatening virus [70]. Foscarnet, a class of antiviral drug approved for the treatment of HIV/AIDS-related cytomegalovirus (CMV) infections and herpes, functions as a pyrophosphate molecule by binding to viral DNA polymerase and preventing the elongation of the DNA chain [71]. *In vitro* studies have revealed that SSYA10–001, a small inhibitor molecule of SARS-CoV helicase, has an antiviral effect on several coronaviruses by likely targeting a conserved binding pocket in nsp13 domain and can lead the production of successful wide spectrum anti-coronavirus drugs including SARS-CoV-2 [72]*.* Scutellarein isolated from *Scutellaria baicalensis* has been historically used in the treatment of inflammation and respiratory diseases, and myricetin present in fruits such as cranberry and vegetables such as *Calamus scipionum* and garlic. Were found to inhibit SARS-CoV helicase (nsp13) via inhibition of ATPase activity being a potential phytochemical inhibitor [73, 74]. Emetine could be re-purposed to treat COVID-19 based on the *in-vitro* antiviral activity against SARS-CoV-2 and its ability to shield chicken embryos from IBV [75].
