3.2. Viral Proteases
The 3-chymotrypsin-like protease (3CLPro) and a papain-like protease (PLPro) function to process NSPs including RdRp, and therefore, inhibition of these proteases should interfere the activity of RdRp (Rathnayake et al., 2020). The antiviral and cell protection efficacy of 3CLPro inhibition has been illustrated in simian Vero cells infected by SARS-CoV-2 (Jin et al., 2020).
Several candidates have been found to have inhibitory effects on 3CLPro. Anti-hepatitis C virus (HCV) drug ravidasvir has the ability to bind and inhibit the 3CLPro of SARS-CoV-2 (Bera, 2021). Similarly, HCV protease inhibitors paritaprevir and simeprevir were also identified as potential inhibitors of SARS-CoV-2 3CLPro (Alamri et al., 2020). Using computational molecular modeling to screen FDA approved drugs and subsequent studying for their inhibitory effects on SARS-CoV-2 3CLpro enzyme in vitro , boceprevir, ombitasvir, paritaprevir, tipranavir, ivermectin, and micafungin were found to exhibit inhibitory effect towards 3CLpro enzymatic activity (Mody et al., 2021).
The efficacy of 3CLPro inhibitors in COVID-19 need to be proved. Lopinavir is a highly potent inhibitor of the human immunodeficiency virus (HIV) protease essential for intracellular HIV assembly. Concomitant oral administration of lopinavir and ritonavir, which blocks the metabolism of lopinavir, increases the anti-viral potency of lopinavir. However, COVID-19 patients receiving the combined treatment lopinavir and ritonavir yielded no significant benefit compared to patients treated with standard-care (Cao et al., 2020a). More specific inhibitors of proteases for SARS-CoV-2 should be evaluated in COVID-19, since a study indicated that lopinavir and ritonavir did not significant inhibit 3ClPro in an in vitro enzymatic assays (Mahdi, Motyan, Szojka, Golda, Miczi & Tozser, 2020). The aforementioned 3CLPro inhibitors may benefit COVID-19 patients, which is needed to be proved in future clinical trials.
In regard of PLPro, activity profiling and crystal structures of inhibitor study indicated that there is a very high level of sequence and structural similarity between SARS-CoV and SARS-CoV-2 PLPro in the substrate binding pocket, suggesting that SARS-CoV PLPro inhibitors can possibly inhibit SARS-CoV-2 PLPro (Rut et al., 2020). Non-covalent small molecule SARS-CoV PLPro inhibitors has been shown to inhibit SARS-CoV-2 PLPro and display antiviral activity in a SARS-CoV-2 infection model (Klemm et al., 2020). Biochemical, structural, and functional characterization investigation revealed that SARS-CoV and SARS-CoV-2 PLPro share 83% sequence identity but exhibit different host substrate preferences. SARS-CoV-2 PLPro preferentially bind to the ubiquitin-like interferon-stimulated gene 15 protein (ISG15), cleaving ISG15 from interferon responsive factor 3 (IRF3) and attenuating type I interferon responses. Whereas SARS-CoV PLPro predominantly targets ubiquitin chains. Inhibition of SARS-CoV-2 PLPro impairs the virus-induced cytopathogenic effect, maintains the antiviral interferon pathway and reduces viral replication in infected cells (Shin et al., 2020).